CN114807152A - Engineered viral vectors reduce induction of inflammation and immune responses - Google Patents

Abstract

The modified viral genome is capable of reducing the induction of inflammation and immune antiviral responses. This is manifested in reduced NF-kB activity, increased viral transduction rates and increased transgene expression. The viral genome is modified by integration of one or more oligonucleotide sequences capable of binding to TLR9 without inducing activation thereof. The oligonucleotide sequence may be synthetic, bacterial, human or from any other source.

Description

Engineered viral vectors reduce induction of inflammation and immune responses

This application is a divisional application of Chinese Invention Patent Application No. 201780045910.3 with the application date of June 8, 2017 and the invention titled "Engineered viral vector reduces the induction of inflammation and immune response".

This invention was made with government support under HG008525 awarded by the National Institutes of Health. The government has certain rights in this invention.

Relevant application data

This application claims priority to US Provisional Application No. 62/347,302, filed June 8, 2016, which is hereby incorporated by reference in its entirety for all purposes.

technical field

The present invention relates to the field of virus-based therapy. In particular, it involves recombinant viruses.

Technical Field of the Invention

Gene therapy has great potential to prevent, treat and cure many human diseases [1]. In 2012, Glybera ^™ (alipogene tiparvovec) became the first viral gene therapy approved for use in the Western world [2]. Glybera ^TM utilizes adeno-associated virus (AAV) as a viral vector to deliver the human lipoprotein lipase (LPL) gene to the muscle cells of patients with LPL deficiency [3]. Numerous AAV clinical trials are currently underway or planned in the United States and the European Union [4-6].

AAV is a small, non-enveloped virus that packages a single-stranded linear DNA genome about 5 kb in length and has been adapted for use as a gene transfer vector [4]. The coding region of AAV is flanked by inverted terminal repeats (ITRs), which serve as the origin of DNA replication and as the primary packaging signal [7,8]. Both positive and negative strands are equally well packaged into virions and capable of infection [9-11]. Additionally, a small deletion in one of the two ITRs allows packaging of self-complementary vectors in which the genome self-anneals after viral uncoating. This results in more efficient transduction of cells, but with half the coding capacity [12,13]. Although AAV is not associated with disease in any person, >70% of people are seropositive for one or most serotypes [14,15]. Typical routes of administration of AAV vectors include intravenous, intramuscular, subretinal and intracranial injection. AAV gene therapy is most commonly used to deliver wild-type genes to treat monogenic diseases, and AAV vectors have been used to transduce cells in the liver, skeletal and cardiac muscle, retina, and central nervous system [16-26]. Additionally, there is now growing interest in using AAVs to deliver CRISPR-Cas9 gene editing or to deliver broadly neutralizing antibodies against infectious diseases such as human immunodeficiency virus (HIV) and influenza A virus [27-32].

Despite some progress in gene therapy, the key issue is the inflammatory response elicited by the viral vector [33,34]. This is best illustrated by a case of high public concern in 1999, when Jesse Gelsinger died of excessive inflammation after 4 days of gene therapy using adenovirus in a clinical trial [35]. Although AAV has been shown to cause much weaker inflammation compared to adenovirus [36,37], Glybera ^™ therapy still includes 12 weeks of immunosuppression, starting three days before Glybera ^™ administration [38]. These immunosuppressive and anti-inflammatory drugs (cyclosporine A, mycophenolate mofetil, and methylprednisolone) weaken the patient's immune system during treatment, and all patients still develop neutralizing antibodies against the AAV capsid, thereby Future reapplications are avoided. Furthermore, many AAV gene therapy clinical trials do not use immunosuppression prophylactically and administer corticosteroids only when signs of inflammation or tissue damage are present, which is associated with variable treatment efficacy [22,23]. Interestingly, consistent with their ability to elicit inflammatory and immune responses, AAV vectors have also been developed as vaccine vectors against infectious diseases and cancer [39-41].

Inflammation has been implicated as a key determinant of successful AAV-mediated transgene expression. It was found that artificial induction of systemic inflammation, such as upregulation of tumor necrosis factor (TNF), resulted in a decrease in transgene expression in the liver following administration of AAV in mice [42]. This suggests that immune tolerance to AAV-encoded transgenes can be disrupted if given an adequate inflammatory response. Another study characterized inflammation in the liver of mice following AAV infection and found a transient increase in liver enzymes in serum, and liver pathology was also consistent with portal and lobular inflammation [43]. Surprisingly, the authors observed that use of the AAVrh.32.33 virus (which induces higher liver enzymes compared to the other AAV serotypes tested) also resulted in a drop in transgene expression below detection levels, again indicating an inflammatory and immune response associated with poor transgene expression.

In hemophilia B clinical trials, a subgroup of patients has been observed to show elevations in liver enzymes, and this transient transaminitis is accompanied by decreased levels of the transgene-encoded factor IX [22,23]. Patients with elevated transaminases were given tapered corticosteroid therapy, which subsequently normalized transaminases levels in serum and rescued further decreases in factor IX expression. Taken together, these observations are compatible with immune-mediated destruction of AAV-transduced hepatocytes and demonstrate that the inflammatory and immune responses elicited by AAV administration are a safety concern and can hinder therapeutic efficacy in humans. Therefore, it would be advantageous to develop viral vectors that essentially avoid triggering inflammation. Furthermore, instead of systemic immunosuppression with drugs, it would be beneficial to avoid triggering a specific immune response.

It has been previously shown that the DNA genome of AAV is sensed by Toll-like receptor 9 (TLR9) during AAV entry into cells through the endocytic pathway [36,44]. TLR9 is a pattern recognition receptor (PRR) found on the endosomal membranes of immune cells such as B cells, monocytes, macrophages, and plasmacytoid dendritic cells, and is associated with unmethylated receptors found in the AAV genome Substrate CpG motif binding [45,46]. This leads to TLR9 dimerization, which initiates a signaling cascade that activates NF-κB (also known as the p52-RelA complex) and induces type I interferons (IFNs). NF-κB in turn drives the transcriptional upregulation of multiple proinflammatory cytokines, such as TNF, leading to inflammation and immune cell recruitment, while secreted IFN induces the expression of many interferon-stimulated genes (ISGs) and establishes an antiviral state. Importantly, genetic ablation of TLR9 in mice abolished the induction of inflammatory cytokines in the liver following AAV treatment and also reduced the formation of antibodies and T cells against AAV [36]. Thus, TLR9 plays a key role in stimulating early inflammatory and innate immune responses during AAV infection, which also contributes to the initiation of adaptive immunity. Finally, two other pattern recognition receptors, TLR2 and TLR4, are involved in eliciting responses to AAV structural proteins [47,48].

In the field of TLR9, a common tool for blocking TLR9 activation in cell culture is short single-stranded DNA oligonucleotides, which bind TLR9 but do not activate it [49,50]. Several such sequences are known to exist - some synthetic, others derived from organisms - and they generally do not share sequence homology [51-59]. Structural studies revealed how inhibitory oligonucleotides tightly bind TLR9 but do not trigger TLR9 dimerization, which is required for TLR9 activation and downstream signaling [60]. In addition to directly binding TLR9 to antagonize its activation, other mechanisms to block TLR9 activation or TLR9-mediated inflammation have been postulated or shown for other TLR9-inhibiting oligonucleotides [reviewed in 49]. These mechanisms include competing receptor-mediated endocytosis or phagocytosis, inhibition of TLR9 trafficking or inhibition of TLR9 processing into functionally active products, inhibition of endosomal acidification or the activity of key proteases in the endosome, or blocking signaling proteins downstream of TLR9 . When these inhibitory oligonucleotides are provided in cell culture medium in trans with TLR9 ligands, such as DNA viruses or CpG-containing oligonucleotides, they are endocytosed and can bind to TLR9, preventing its stimulatory ligand activation. Replenishment of inhibitory oligonucleotides in trans is widely used in immunology experiments, but it is unclear in the art whether integrating these sequences into the viral genome can prevent it from triggering inflammatory and immune responses.

Although AAV has been shown to elicit a much weaker inflammatory response compared to adenovirus, Glybera ^™ tepaliprkin treatment still included 12 weeks of immunosuppression starting 3 days prior to Glybera ^™ tepaliprkin administration. These immunosuppressive drugs strongly block T cell activation, thus weakening the patient's immune system during treatment. It would be advantageous to design viral vectors that avoid and cause a reduced or no inflammatory response when administered. Furthermore, it would be beneficial if the immunosuppression was not systemic and if it was transient. Prevention of inflammation and immune responses can also improve transgene expression and allow re-administration of viral vectors for future purposes.

There is a continuing need in the art to improve the efficacy of viral vectors for use in therapy and in vivo production of biological products.

SUMMARY OF THE INVENTION

According to one aspect of the present invention, nucleic acid molecules are provided. It comprises the viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation.

According to another aspect, recombinant viruses for delivering desired functions to mammalian cells are provided. The recombinant virus comprises a viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation.

Another embodiment is a method of treating a mammal. The method includes administering the recombinant virus to a mammal in need thereof. The recombinant virus comprises a viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation.

Another aspect is a method of making a viral genome of a recombinant virus. Inhibitory nucleic acid sequences are inserted into the viral genome. The inhibitory nucleic acid sequence binds to TLR9 but does not trigger TLR9 activation.

According to one aspect, nucleic acid molecules are provided. It contains inverted terminal repeats (ITRs) and nucleic acid sequences that inhibit TLR9-mediated inflammation.

Another aspect of the invention is a nucleic acid molecule. The molecule comprises a viral genome covalently linked to an inhibitory nucleic acid sequence that inhibits TLR9-mediated inflammation.

Another aspect of the present invention is a recombinant virus for delivering a desired function to mammalian cells. The recombinant virus comprises a viral genome comprising inhibitory nucleic acid sequences that inhibit TLR9-mediated inflammation.

Another aspect of the invention is a method of making a viral genome of a recombinant virus. The nucleic acid sequence is inserted into the viral genome. The nucleic acid sequence inhibits TLR9-mediated inflammation.

According to another aspect of the present invention, nucleic acid vectors are provided. The vector comprises at least one nucleic acid sequence. The nucleic acid sequence is capable of inhibiting TLR9-mediated inflammation.

Another aspect of the invention is a method of reducing the immunogenicity of a genome-modified virus. The method includes inserting a nucleic acid sequence into the genome. The nucleic acid sequence inhibits TLR9-mediated inflammation. The modified virus elicits a reduced inflammatory response in the host compared to the virus without the inhibitory sequence.

Another aspect of the invention is a method of increasing expression in a host cell of a virally introduced transgene. The method involves introducing into a host a modified virus having a genome. The genome comprises nucleic acid sequences. The nucleic acid sequence inhibits TLR9-mediated inflammation. The modified virus results in higher transgene expression in host cells than viruses without inhibitory sequences.

Another aspect of the invention is a composition comprising a viral capsid that encapsulates a nucleic acid sequence that inhibits TLR9-mediated inflammation.

These and other aspects and embodiments, which will become apparent to those of skill in the art upon reading this specification, provide the art for better use of viral vectors and virions for better treatment of mammals and for better use of viral vectors and virions in Tools for the production of transgenic products in host cells, host tissues and host animals.

Any and all of the above aspects can be combined with any of the following features.

The viral genome may be an adeno-associated virus (AAV) genome.

The viral genome can be selected from adenovirus, herpes simplex virus, varicella, variola virus, hepatitis B virus, cytomegalovirus, JC polyoma virus, BK polyoma virus, monkeypox virus, herpes zoster virus, Epstein-Barr virus, human herpesvirus 7, Kaposi's sarcoma-associated herpesvirus, and human parvovirus B19.

The viral genome can be single-stranded.

Viral genomes can be packaged in virions.

The viral genome can contain genes that can be expressed in human cells.

The viral genome can be a cytotoxic virus used to lyse target tumor cells.

The inhibitory nucleic acid sequence may comprise the c41 oligonucleotide sequence TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1).

The inhibitory nucleic acid sequence may comprise multiple copies of the c41 sequence (SEQ ID NO: 1).

The inhibitory nucleic acid sequence may comprise two copies of the c41 sequence (SEQ ID NO: 1) separated by a linker sequence.

The linker sequence is AAAAA (SEQ ID NO:8).

The inhibitory nucleic acid sequence can be selected from:

ODN 2088: TCC TGG CGG GGA AGT (SEQ ID NO: 2);

ODN 4084-F: CCTGGATGGGAA (SEQ ID NO: 3);

ODN INH-1: CCTGGATGGGAATTCCCATCCAGG (SEQ ID NO: 4);

ODN INH-18: CCT GGA TGG GAA CTT ACC GCT GCA (SEQ ID NO: 5);

ODN TTAGGG:TT AGG GTT AGG GTT AGG GTT AGG G (SEQ ID NO: 6); and

G-ODN: CTC CTA TTG GGG GTT TCC TAT (SEQ ID NO: 7).

The inhibitory nucleic acid sequence can be a bacterial sequence.

The viral genome, recombinant virus, vector or nucleic acid sequence may contain non-human genes.

Inhibitory nucleic acid sequences can be inserted downstream or within the 3' untranslated region of the viral genome.

The viral genome can be covalently linked to the inhibitory nucleic acid sequence through phosphodiester bonds.

The viral genome, recombinant virus, vector or nucleic acid sequence may contain a detectable marker.

The detectable label can be inducible.

The inhibitory nucleic acid sequence may comprise the human telomeric sequence set forth in SEQ ID NO:9.

Viral genomes can be self-complementary.

The viral genome can be covalently linked to a variety of inhibitory nucleic acid sequences.

Various inhibitory nucleic acid sequences can include inhibitory sequences and their reverse complements.

The inhibitory nucleic acid sequence may comprise three copies of the c41 sequence (SEQ ID NO: 1), each copy being separated by a linker sequence.

The inhibitory nucleic acid sequence can be selected from:

ODN 2114: TCCTGGAGGGGAAGT (SEQ ID NO: 16);

ODN 4024: TCCTGGATGGGAAGT (SEQ ID NO: 17);

ODN INH-4: TTCCCATCCAGGCCTGGATGGGAA (SEQ ID NO: 18);

ODN INH-13: CTTACCGCTGCACCTGGATGGGAA (SEQ ID NO: 19);

ODN Poly-G: GGGGGGGGGGGGGGGGGGGG (SEQ ID NO: 20);

ODN GpG: TGACTGTGAAGGTTAGAGATGA (SEQ ID NO: 21);

ODN IRS-869: TCCTGGAGGGGTTGT (SEQ ID NO: 22);

ODN IRS-954:TGCTCCTGGAGGGGTTGT (SEQ ID NO:23); and

ODN 21158: CCTGGCGGGG (SEQ ID NO: 24).

The inhibitory nucleic acid sequence can be ODN TTAGGG (SEQ ID NO: 6).

The inhibitory sequence can be covalently linked to the linker.

The inhibitory sequence can be upstream of the linker.

The inhibitory nucleic acid sequence may comprise multiple copies of ODN TTAGGG (SEQ ID NO: 6).

Multiple copies of ODN TTAGGG (SEQ ID NO: 6) can each be separated by a linker.

The inhibitory nucleic acid sequence may comprise at least 2, at least 3, at least 4, or at least 5 copies of ODN TTAGGG (SEQ ID NO: 6), each copy being separated by a linker.

The inhibitory nucleic acid sequence can be a human sequence.

The viral genome, recombinant virus, vector or nucleic acid sequence may comprise non-human nucleic acid sequences.

The viral genome, recombinant virus, vector or nucleic acid sequence may comprise human genes.

The viral genome, recombinant virus, vector or nucleic acid sequence may comprise human nucleic acid sequences.

Inhibitory nucleic acid sequences can be inserted in the 5' untranslated region of the viral genome.

Inhibitory nucleic acid sequences can be inserted upstream of the promoter in the viral genome.

The viral genome, recombinant virus, vector or nucleic acid sequence may contain an inducible promoter.

The inhibitory nucleic acid sequence may comprise two repeat monomers of SEQ ID NO:1.

The inhibitory nucleic acid sequence may comprise the three repeat monomers of SEQ ID NO:1.

The inhibitory nucleic acid sequence may comprise SEQ ID NO:6 or SEQ ID NO:9.

The inhibitory nucleic acid sequence may comprise the three repeat monomers of SEQ ID NO:6 or SEQ ID NO:9.

The inhibitory nucleic acid sequence may comprise the 5 repeat monomers of SEQ ID NO:6 or SEQ ID NO:9.

The application steps can be repeated.

Viral genomes can be packaged in virions.

The insertion step can use DNA ligase.

The viral genome can be single-stranded when in a virion.

The viral genome of a recombinant virus can contain genes for delivery to and expression in human cells.

The viral genome, recombinant virus, vector or nucleic acid sequence may contain linkers that separate each such nucleic acid sequence.

The viral genome, recombinant virus, vector or nucleic acid sequence may comprise at least 2, at least 3, at least 4, or at least 5 copies of the nucleic acid sequence.

The inhibitory nucleic acid can be covalently linked to the gene.

The inhibitory nucleic acid sequence may be 95% identical to the c41 oligonucleotide sequence TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1).

The inhibitory nucleic acid sequence can be 95% identical to SEQ ID NO:9.

The inhibitory nucleic acid sequence may be 95% identical to a sequence selected from:

ODN 2088: TCC TGG CGG GGA AGT (SEQ ID NO: 2);

ODN 4084-F: CCTGGATGGGAA (SEQ ID NO: 3);

ODN INH-1: CCTGGATGGGAATTCCCATCCAGG (SEQ ID NO: 4);

ODN INH-18: CCT GGA TGG GAA CTT ACC GCT GCA (SEQ ID NO: 5);

ODN TTAGGG: TT AGG GTT AGG GTT AGG GTT AGG G (SEQ ID NO: 6); and

G-ODN: CTC CTA TTG GGG GTT TCC TAT (SEQ ID NO: 7). The inhibitory nucleic acid sequence may be 95% identical to a sequence selected from:

ODN 2114: TCCTGGAGGGGAAGT (SEQ ID NO: 16);

ODN 4024: TCCTGGATGGGAAGT (SEQ ID NO: 17);

ODN INH-4: TTCCCATCCAGGCCTGGATGGGAA (SEQ ID NO: 18);

ODN INH-13: CTTACCGCTGCACCTGGATGGGAA (SEQ ID NO: 19);

ODN Poly-G: GGGGGGGGGGGGGGGGGGGG (SEQ ID NO: 20);

ODN GpG: TGACTGTGAAGGTTAGAGATGA (SEQ ID NO: 21);

ODN IRS-869: TCCTGGAGGGGTTGT (SEQ ID NO: 22);

ODN IRS-954: TGCTCCTGGAGGGGTTGT (SEQ ID NO: 23); and

ODN 21158: CCTGGCGGGG (SEQ ID NO: 24).

The inhibitory nucleic acid sequence may comprise multiple copies of SEQ ID NO:6 and/or SEQ ID NO:9.

The inhibitory nucleic acid sequence may comprise two copies of SEQ ID NO:6 and/or SEQ ID NO:9 separated by a linker sequence.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1. Schematic representation of viral gene therapy and inflammatory and immune responses.

Figure 2. Schematic showing TLR9 sensing of AAV DNA during viral entry and inflammatory and immune responses. From Rogers et al., 2011, Frontiers in Microbiology, "Innate Immune Responses to AAVVectors," Vol. 2, Paper 194.

Figures 3A-3B. Figure 3A shows the nucleotide sequence of single stranded oligonucleotide c41 (SEQ ID NO: 1). Figure 3B shows the organization of AAV-eGFP and AAV-eGFP-c41 genomes. ITR, inverted terminal repeat; LPA, late polyadenylation signal. The target transgene described in this case is eGFP. The insert has the c41 sequence. The spacer depicted in Figure 3B is shown in SEQ ID NO:8.

Figure 4 shows NF-κB activity in HEK293 TLR9 cells mock-infected or infected with AAV virus (DJ capsid, self-complementary AAV genome, encoding eGFP). **, p<0.005; n.s., not significant. Crude virus preparations were used for this experiment.

Figure 5. Flow cytometry histogram showing GFP expression in HEK293 TLR9 cells mock-infected or infected with AAV virus with or without c41. Crude virus preparations were used for this experiment.

Figures 6A-6B. Figure 6A shows the nucleotide sequence of "Telomere" (SEQ ID NO:9), which is a single nucleotide sequence containing a (TTAGGG) ₄ (SEQ ID NO:6) motif from mammalian telomeres chain oligonucleotides. Figure 6B shows the organization of the AAV-eGFP-telomere genome. Inserts have "telomere" sequences. The spacer shown in Figure 6B is SEQ ID NO:8.

Figures 7A-7B. Figure 7A shows the percentage of transduced cells (GFP+) 2 days after infection of B cell lines with similar amounts of the indicated AAV viruses, analyzed by flow cytometry. Figure 7B shows TNF production in primary human CD14+ monocyte supernatants at 18 hours post infection as determined by ELISA.

Figures 8A-8B. Design of self-complementary AAV vectors. (Fig. 8A) DNA sequences of "c41" and "telomere". (FIG. 8B) Genomic organization of AAV vector (scAAV-eGFP) and modified vector. LpA: polyA signal.

Figures 9A-9C. Inflammatory responses to various AAV vectors in human immune cells in vitro. ( FIG. 9A ) Primary human macrophages were infected with AAV2 virus (MOI: 10 ⁵ vg/cell), and the supernatant was collected 18 hours later and analyzed for TNF levels by ELISA. 5 μM ODN2006, a CpG-containing oligonucleotide, was used as a positive control. (FIG. 9B, FIG. 9C) Primary human CD14+ monocytes from two different donors were infected and analyzed for TNF levels similarly to (A). Data shown are the mean±sd of n=3 technical replicates. *P<0.05 (unpaired t-test) compared to scAAV-eGFP.

Figures 10A-10E. Further characterization of AAV vectors. (FIG. 10A) "Control" DNA sequence. (FIG. 10B) Genomic organization of modified vectors. (FIG. 10C) Primary human macrophages were infected with AAV2 virus similarly to (FIG. 9A) and analyzed for TNF levels by ELISA. Primary human monocytes were infected (Fig. 10D) similarly to (Fig. 9B, Fig. 9C) and analyzed for TNF levels by ELISA. Data shown (Fig. 10C, Fig. 10D) are the mean ± s.d. of n=3 technical replicates. *P<0.05 (unpaired t-test) compared to scAAV-eGFP. (Fig. 10E) Adult C57BL/6 mice were infected with the indicated AAV2 viruses similarly to (Fig. 12A, Fig. 12B and Fig. 12C) and a piece of liver was analyzed by qRT-PCR for indicated gene expression. Data shown are the mean ± s.d. of n=5 mice per condition (except n=3 mice for the scAAV-eGFP-3x control). *P<0.05 (unpaired t-test) compared to saline conditions. N.s.: Not significant (P>0.05).

Figures 11A-11B. (Figure 11A) Primary human macrophages were infected with two different batches of AAV2 viruses similarly to (Figure 9A) and analyzed for TNF levels. (FIG. 11B) HeLa cells were infected with AAV2 virus at the indicated MOI, cells were harvested 48 hours later, and their GFP expression was analyzed by flow cytometry. The percentage of GFP positive cells is shown. Data shown are mean±s.d. of n=3 technical replicates. *P<0.05 (unpaired t-test) compared to scAAV-eGFP.

Figures 12A-12C. Inflammatory responses to intravenous administration of various AAV vectors in adult mice in vivo. (Fig. 12A, Fig. 12B and Fig. 12C) Adult C57BL/6 mice were infected with the indicated AAV2 viruses ( ¹⁰¹¹ vg/mouse) by tail vein injection. After 2 hours, animals were euthanized and a piece of liver was analyzed by qRT-PCR for indicated gene expression. For each gene, saline injection was set to 1-fold expression. Data shown are the mean±sd of n=3 mice per condition (FIG. 12A) or n=4 mice per condition (FIGS. 12B and 12C). *P<0.05 (unpaired t-test)*, compared to saline conditions. Ns: not significant (P>0.05).

Figure 13. Genomic organization of single-stranded AAV vectors (ssAAV-eGFP) and ssAAV-eGFP-5x telomeres.

Figures 14A-14C. Inflammatory and immune responses in neonatal mice following subretinal administration of various AAV vectors. (FIG. 14A, FIG. 14B and FIG. 14C) Neonatal CD1 mice (P1) received the indicated AAV8 viruses (1.8x10&lt; ⁸ &gt; vg/mouse eye) by subretinal injection. At P21, animals were euthanized and the eye cups were dissected out. The rest of the retina and eye cup were analyzed for expression of the indicated genes by qRT-PCR. For each gene, saline injection was set to 1-fold expression. Each triangle represents an animal. Data shown are for n=3 mice (saline) and n=5 mice (each virus) and mean values are shown.

Figures 15A-15C. Analysis of immune cell markers in the retina of neonatal mice in vivo following subretinal administration of various AAV vectors. (FIG. 15A, FIG. 15B and FIG. 15C) Similar to (FIGS. 14A-14C), the retina was analyzed for the expression of the indicated genes by qRT-PCR. Aif1 (Iba1) is known to be expressed in microglia, while Cd4 and Cd8a are markers for helper and cytolytic T cells, respectively. For each gene, saline injection was set to 1-fold expression. Each triangle represents an animal. Data shown are for n=3 mice (saline) and n=5 mice (each virus) and mean values are shown.

Figure 16. GFP expression in the eye. Neonatal CD1 mice (P1) received the indicated AAV8 virus ( ^1.8x108 vg/mouse eye) by subretinal injection. At P30, animals were euthanized and GFP expression was observed in flat-mounted eye cups. Data shown are n=2 mice per condition.

Figures 17A-17B. Figure 17A shows the nucleotide sequence of c41 (SEQ ID NO: 1), which is a single-stranded oligonucleotide. Figure 17B shows the organization of AAV-eGFP and AAV-eGFP-c41 genomes. ITR, inverted terminal repeat; LPA, late polyadenylation signal. The target transgene described in this case is eGFP. The insert has the c41 sequence. The spacer depicted in Figure 17B is shown in SEQ ID NO:8.

Figures 18A-18B. Figure 18A shows the nucleotide sequence of "Telomere" (SEQ ID NO:9), which is a single stranded containing (TTAGGG) ₄ (SEQ ID NO:6) motif from mammalian telomeres Oligonucleotides. Figure 18B shows the organization of the AAV-eGFP-telomere genome. Inserts have "telomere" sequences. The spacer shown in Figure 18B is SEQ ID NO:8.

Detailed description of the invention

The inventors have developed viral vectors and virions that have their own protective effects against the host immune and inflammatory systems. These vectors and virions carry short nucleic acid sequences that inhibit the activation of toll-like receptor 9 (TLR9), a host protein that activates inflammatory and immune responses in mammalian cells.

The short nucleotide sequence used to inhibit TLR9 can be of any origin. It can be of bacterial, human, synthetic or other origin. One particular sequence is the 20 nucleotide "c41" from Pseudomonas aeruginosa [TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1)]. Another specific sequence is derived from human telomeres and contains (TTAGGG) ₄ (SEQ ID NO:6). Additional inhibitory sequences are shown in SEQ ID NOs: 2-5, 7, 9 and 16-24. Inhibitory sequences with at least 80% homology/identity to these sequences can also be used. Inhibitory sequences that share at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% and/or at least 99% homology/identity with these sequences can also be used. Multiple copies of the inhibitory sequences can be used in tandem array format or in the form of separated in a viral vector by spacer or linker sequences or other parts of the viral genome. In some embodiments, one, two, three, four, five, six, seven, eight, nine, ten, fifteen or twenty copies are used. In some embodiments, one or more copies of the inhibitory sequence are on the plus strand and some copies are on the minus strand of the viral genome.

The inhibitory oligonucleotide sequence is introduced into the host cell as part of the viral genome or virion rather than as a separate agent. This makes the effect of the oligonucleotide sequence local rather than systemic. Furthermore, immune evasion is a transient phenomenon as it occurs during AAV or other viral entry, unlike immunosuppression with drugs that can last for weeks. Additionally, it ensures that the beneficial antagonist activity occurs where it is needed - using a virus or viral genome. If the virus or viral genome is not replicating in the host cell, the effect of the oligonucleotide will be short-lived. If the virus or viral genome replicates, the effect will be co-expanded with the replication.

Inhibitory nucleic acid sequences can be inserted into the viral genome using any recombinant DNA engineering method. This can involve in vitro or in vivo recombination. For example, in vitro recombination can be accomplished using DNA ligases or other nucleic acid ligases. In vivo recombination can be achieved by co-transforming host cells with separate donor molecules that share the homology by which they will recombine using the host cell machinery. Alternatively, a single donor molecule can be recombined with host cell sequences in vivo. Combinations of these methods can also be used. Typically, insertion will involve the standard linkage (phosphodiester bond) of one deoxyribonucleotide to another deoxyribonucleotide. However, there may be situations where non-standard linkages would be used between the inhibitory nucleic acid sequence and the rest of the viral genome. Optionally, the inhibitory nucleic acid sequence is located in an untranslated region of the viral genome.

The genome may optionally contain therapeutic genes and/or marker genes. Typically, the gene is a non-viral gene, or a gene that is not naturally present in the viral genome. The gene is capable of being expressed in mammalian host cells or animals. Expression can be under the control of a viral promoter or a promoter introduced with the gene. As an example, expression can be inducible, repressible, condition-responsive or constitutive. Therapeutic genes are genes that encode RNA or protein products that are beneficial to the host. For example, the benefit can be improved health, prevention of infection, or remediation of deficiencies. Labels allow one to track the location, level of replication, level of proliferation, level of transcription or translation of the virus or its products or components. Suitable labels include readily detectable labels such as fluorescent proteins, chromoproteins, and the like. Optionally, a second reagent can be used or added for detection of labeled proteins or for visualization of detectable substances. The introduced gene can be human or non-human, heterologous (from another species) or homologous (from the same species) or endogenous (from the same subject).

Any DNA viral genome can be used, whether single-stranded or double-stranded. Examples of suitable viruses that may be used include, but are not limited to, adeno-associated virus (AAV), adenovirus, herpes simplex virus, chickenpox, smallpox virus, hepatitis B virus, cytomegalovirus, JC polyoma virus, BK polyoma virus, monkey Wild-type or variants of poxvirus, herpes zoster virus, Epstein-Barr virus, human herpesvirus 7, Kaposi's sarcoma-associated herpesvirus, human parvovirus B19, and enterovirus. Viruses can be, but are not limited to, cytotoxic, cytolytic, or cause latent infection. Viral vectors in which the viral genome has been modified can also be used. For example, genomes modified to encode fewer viral proteins can be used. As another example, viral genomes modified to not encode viral proteins can be used. As a non-limiting example, the viral genome can include inverted terminal repeats and/or other non-coding genetic elements that facilitate packaging of the engineered viral genome into the capsid.

The viral genome can be delivered to mammalian host cells as naked DNA, in liposomes, complexed with polymers, in a concentrated or compacted state, as gold particles, as virions, or in any other manner suitable for administration. Usually the complete viral genome will be administered, but in some cases it may be necessary to use a partial genome. Parts of the genome may be complemented by helper functions provided by the host cell or by another genomic or viral entity. For example, partial genomes can be used if the therapeutic payload is large and some essential viral functions must be omitted for packaging.

The recombinant virus can be administered to the mammal or mammalian cells according to any route effective for this purpose. Administration can be systemic, eg, by blood. It can be delivered orally, subcutaneously, topically, bucally, anally, intramuscularly, intravenously, intratumorally, intracranically, intrathecally, subretinal, and the like. Any suitable carrier or vehicle can also be used for administration. It may be necessary to pretreat cells or mammals to make them more permeable or receptive to the recombinant virus. A mammal "in need" of a recombinant virus can be a mammal for which the virus is beneficial. It may be a mammal suffering from a disease or defect. It can be the mammal to be diagnosed or analyzed. Even if it does not have a disease or defect, it can be a mammal that can benefit from the administered recombinant virus.

Taken together, our results show that the integration of c41 or human telomeric sequences into the AAV genome (a) does not reduce viral packaging and infectivity, (b) prevents TLR9-mediated inflammation, and (c) reduces pro-inflammatory cytokines induction of and (d) increased transgene expression. The increased transgene expression can be attributed to a reduced immune response, as TLR9 activation also induces interferon expression, which triggers an antiviral state. The engineered immune evasion properties we show below are specific (against TLR9), transient (eg, can occur during viral entry), and do not result in systemic immunosuppression (against AAV-infected immune cells only).

The inhibitory nucleic acid sequences we used, as well as other viruses known in the art, can be incorporated into other viruses that, like AAV, have potential utility in humans and other mammals, but elicit potentially undesirable inflammation/immunity reaction. For example, oncolytic viruses that preferentially infect and lyse cancer cells are used to kill or shrink tumors. These viruses are replicative (unlike AAV vectors used in gene therapy), so they can release new virions to shrink remaining tumors. Examples include wild-type or variants of herpes simplex virus, adenovirus, and enterovirus. Several reports have shown that immunosuppression produced by chemotherapy can enhance oncolytic virus therapy because the immune system normally tries to inactivate the oncolytic virus, preventing it from infecting cancer cells. Therefore, integration of inhibitory oligonucleotides into the genome of oncolytic viruses such as herpes simplex virus has the potential to allow them to evade immune clearance and persist for longer periods of time for oncolysis.

The above disclosure generally describes the present invention. All references disclosed herein are expressly incorporated by reference. A more complete understanding can be obtained by reference to the following specific examples, which are provided for illustrative purposes only and are not intended to limit the scope of the present invention. The disclosure of the present invention includes all embodiments expressly recited in the claims. Furthermore, all features disclosed in the dependent claims apply equally to the independent claims on which they are based on the other independent claims. Accordingly, such combinations of dependent claims with other independent claims are expressly contemplated and disclosed.

Example 1 - Construction of modified viral genomes

To design an AAV vector with the ability to specifically evade TLR9 activation in immune cells, we inserted two copies of c41 separated by a 5-nucleotide-long spacer (AAAAA; SEQ ID NO: 8) in a gene encoding enhancer The 3' untranslated region of the AAV vector of green fluorescent protein (eGFP) (Fig. 3B). Subsequently, we produced wild-type AAV-eGFP virus and AAV-eGFP-c41 virus (containing two c41 inserts). The infectious titers of the two viruses were comparable (~ ¹⁰⁹ infectious units/ml, as determined by titration on HeLa cells), indicating that the addition of c41 to the viral genome does not impede viral packaging and infectivity, a requirement for large-scale Important considerations for the generation of viral vectors for gene therapy.

Example 2 - Modified viral genome reduces NF-κB activation

To measure the inflammatory response, we used HEK293 cells stably expressing TLR9 (HEK293 TLR9 cells), which senses the AAV DNA genome and also expresses alkaline phosphatase under the transcriptional control of NF-kB. When NF-κB is activated, which is indicative of inflammation, alkaline phosphatase is secreted into the medium and acts on the provided substrate, resulting in a change in the color of the medium that can be measured on a plate reader. We mock infected HEK293TLR9 cells or infected them with AAV-eGFP or AAV-eGFP-c41. Consistent with the literature, AAV-eGFP infection induced a small but statistically significant increase in NF-κB activity (Figure 4). In contrast, AAV-eGFP-c41 infection was not significantly different compared with mock-infected cells, suggesting that the virus is able to evade initiating an inflammatory response.

Example 3 - Modified viral genomes transduce more cells and express more transgenes

We analyzed the above three conditions for eGFP expression (depicted in Figure 4) using flow cytometry. We found that AAV-eGFP-c41 transduced more cells than AAV-eGFP (52.7% GFP+ vs 34.6% GFP+) (Figure 5). Furthermore, eGFP from AAV-eGFP-c41-infected GFP+ cells expressed approximately twice as much eGFP as from AAV-eGFP-infected GFP+ cells (mean fluorescence intensity [MFI] 5335 vs. 2749).

In conclusion, we engineered AAV vectors to escape TLR9-mediated inflammation by integrating inhibitory oligonucleotides into the viral genome.

Example 4 - Integration of c41 or telomeric sequences in the AAV genome transduced more cells and reduced TNF induction.

We inserted three copies of a "telomere" (a sequence derived from mammalian telomeres that contains the inhibitory (TTAGGG) ₄ motif (SEQ ID NO:6), which has been shown to block TLR9 signaling (Fig. 6A) and Figure 6B)). When titrated on HeLa cells, AAV-eGFP-telomere virus produced similar viral titers to AAV-eGFP and AAV-eGFP-c41, demonstrating that integration of 'telomeres' does not hinder viral packaging and infectivity.

When we infected B cell lines with similar amounts of AAV-eGFP or AAV-eGFP-c41 or AAV-eGFP-telomere, both AAV-eGFP-c41 and AAV-eGFP-telomere were more transduced than AAV-eGFP more cells (Figure 7A). This finding also suggests that integration of inhibitory sequences in the AAV genome increases transgene expression.

Subsequently, we harvested primary human CD14+ monocytes from blood and subjected them to infection conditions similar to those described above. We performed ELISA on the supernatant to analyze the production of TNF, as TNF is the prototypical pro-inflammatory cytokine induced by NF-κB activation. AAV-eGFP infection increased TNF production compared to mock infection, whereas AAV-eGFP-c41 and AAV-eGFP-telomere infections showed no or little increase in TNF production (Figures 7A-7B), suggesting that these two viruses are capable of Escape triggers an inflammatory response.

Example 5 - Design of a self-complementary AAV vector

Researchers typically use short inhibitory oligonucleotides (usually 10-30 nucleotides in length) to antagonize TLR9 signaling in cell culture. However, it is unclear whether these inhibitory oligonucleotides retain functionality in the context of a much larger viral genome (ie, the sequence is covalently linked at both ends to a much longer sequence). To test this possibility, we utilized a self-complementary (sc) AAV vector encoding enhanced green fluorescent protein (eGFP) and 3 copies of "c41" or "telomere" from bacterial and mammalian telomeres, respectively [52,57,58,61] were inserted into plasmids containing the vector genome (Figures 8A and 8B). We started with sc AAV vectors as they have been shown to be more efficient at eliciting TLR9 activation and induce more inflammation in mouse livers than single-chain (ss) AAV vectors. Since 'c41' and 'telomeres' are predicted to have strong secondary structures, we used AAAAA linkers between copies of the inhibitory oligonucleotide. Additionally, the 3xc41 and 3x telomeric sequences are placed after the polyA sequence and upstream of the right inverted terminal repeat (ITR), so they will be present in the DNA genome during viral entry, but not after successful transduction in subsequent mRNA transcripts ("scAAV-eGFP-3xc41" and "scAAV-eGFP-3x telomeres"). Finally, to determine whether the location of the inhibitory oligonucleotide in the viral genome is important, we also generated a vector in which the 3x telomere is located between the left ITR and the promoter ("scAAV-3x telomere-eGFP").

Example 6 - In vitro inflammatory response of primary human macrophages and monocytes

We packaged various AAV vectors into AAV2 serotypes and infected primary human monocyte-derived macrophages at a multiplicity of infection (MOI) of 10 ⁵ viral genomes (vg) per cell. As expected, we found that scAAV-eGFP infection of macrophages caused a strong induction in the supernatant of TNF, a prototypical inflammatory cytokine with a well-defined role in stimulating fever, apoptosis and inflammation and was produced following TLR9 signaling and NF-κB activation (Fig. 9A). In contrast, both scAAV-eGFP-3x41 and scAAV-eGFP-3x telomeres significantly reduced TNF induction by >95%, indicating the integration of "c41" or "telomeres" in these viruses compared to wild-type (WT) vectors Can escape triggering an inflammatory response. In addition, scAAV-3x telomere-eGFP was also able to prevent TNF induction by >95%, demonstrating that inserted inhibitory oligonucleotides can be placed in other parts of the viral genome and retain the ability to block inflammation. Mock infection with phosphate buffered saline (PBS) and treatment with ODN 2006, a commercially available CpG-containing oligonucleotide known to strongly activate TLR9/NF-kB and inflammation, were considered negative, respectively and positive control. We tested primary human CD14+ monocytes and again found that scAAV-eGFP elicited strong TNF induction, whereas scAAV-eGFP-3xc41 and scAAV-eGFP-3x telomeres abolished most TNF induction (Figure 9B). scAAV-3x telomere-eGFP also reduced TNF induction, although the inhibition was -85%, which may be due to differences between cell types or donor tissues. TNF-induced escape was also reproduced in primary CD14+ monocytes obtained from another donor (Figure 9C).

As further characterization, we inserted 3 copies of a "control" (ie random sequence that does not block TLR9) or 1 copy of a "telomere" into the plasmid containing the vector genome (Fig. 10A, Fig. 10B). We chose the sequence "control" as it has been used as a negative control oligonucleotide in TLR9 experiments. We found that scAAV-eGFP-1x telomeres were able to reduce TNF induction compared to scAAV-eGFP in human macrophages, but not as efficiently as scAAV-eGFP-3x telomeres (Fig. 10C). This suggests that 1 copy of telomeres can reduce inflammation. We also observed that the scAAV-eGFP-3x control elicited TNF secretion as efficiently as scAAV-eGFP in human monocytes, suggesting that insertion of a sequence that inhibits TLR9 is required to block inflammation (Fig. 10D).

As AAV vectors are considered biologics and may exhibit batch-to-batch variability, we produced another batch of scAAV-eGFP and scAAV-eGFP-3x telomeric AAV2 viruses and found that scAAV-eGFP-3x telomere was comparable to WT It was able to reduce TNF induction by -75% compared to vehicle (Figure 11A). Based on various viral preparations as well as donor monocytes and macrophages (Figures 9A-9C, Figures 10A-10E and Figures 11A-11B), we concluded that ours contained 3 copies compared to the WT vector The "c41" or "telomere" engineered vector reduces TNF induction by approximately 75-98%. On average, scAAV-eGFP-3x telomeres reduced TNF induction by ~85% compared to scAAV-eGFP. Importantly, we did not observe differences in viral titers (determined by qPCR against the viral genome) obtained from generating any of the above AAV2 vectors, indicating that the engineered vectors were not defective in packaging (data not shown). Furthermore, when we infected HeLa cells (a permissive cell line widely used to titrate AAV infectivity) with scAAV-eGFP and scAAV-eGFP-3x telomeres at a range of MOIs, we did not observe 4+ viral titers A log difference in transduction (% GFP+ cells) demonstrated that the engineered vectors were equally capable of transducing cells (FIG. 11B).

Example 7 - Inflammatory Response in Liver Tissue in Mice

Intravenous delivery of AAV is commonly used to transduce hepatocytes for gene therapy. Previous studies have shown that after intravenous administration of AAV, Kupffer cells (resident liver antigen-presenting cells) in the mouse liver are able to sense the sc AAV genome and initiate inflammation and innate immune responses after 1–9 h [36]. These responses include induction of proinflammatory cytokines such as TNF and IL6 and type I interferons such as IFN-beta. TLR9-/- mice did not exhibit these inflammatory and innate immune responses in the liver, demonstrating the central role of TLR9 as an innate immune sensor in vivo. Additionally, immune cells such as neutrophils, macrophages and natural killer (NK) cells infiltrated the liver 2 hours after AAV administration. To determine whether our engineered vector could reduce inflammation in the liver in vivo, we administered PBS or an equivalent amount of scAAV-eGFP or scAAV-eGFP-3x telomeres via tail vein injection. We chose scAAV-eGFP-3x telomeres for in vivo characterization because "telomeres" are derived from human sequences and may be preferred for clinical applications. Consistent with previous work, scAAV-eGFP stimulated increased expression of Tnf and Il6 in the liver (approximately 3- to 10-fold compared to saline), indicating inflammation (FIG. 12A). In contrast, scAAV-eGFP-3x telomeres showed little or no increase in inflammatory markers. We tested more mice in subsequent experiments and found that compared with saline, scAAV-eGFP stimulated a statistically significant induction of Tnf in liver, whereas scAAV-eGFP-3x telomeres and scAAV-eGFP- 3xc41 did not have this stimulatory effect (Figures 12B and 12C), indicating their ability to evade triggering liver inflammation. Finally, we demonstrated that the scAAV-eGFP-3x control did not prevent inflammation in the liver compared to scAAV-eGFP (Fig. 10E).

Example 8 - Design of single-chain AAV vectors and in vivo determination of inflammatory responses in mouse ocular tissue

Next, we designed the single-stranded AAV vector ssAAV-eGFP by inserting 5 copies of the "telomere" with the AAAAA linker, and then inserting the other 5 copies in the plasmid in antisense orientation, resulting in ssAAV-eGFP-5x Telomeres (Figure 13). Since both the plus and minus strands of the viral genome are equally likely to be packaged into viral particles, this ensures that each packaged viral genome has five copies of "telomeres" in the correct orientation. Two AAV8 viruses were generated and purified. Again, we did not observe a difference in titer between the two vectors, indicating similar packaging efficiencies (data not shown). ssAAV-eGFP was chosen because it has previously been used for subretinal injection in mice and transduces photoreceptors with high efficiency in the eye [62].

Several studies have shown that AAV gene therapy in the eye and brain appears to be generally safe [63]. Although the eye is generally considered a site of immune privilege, it is known to contain microglia, the resident macrophages of the central nervous system, which have been reported to express TLR9 and respond to CpG motifs [64-67]. A recent study delivering AAV vectors in cynomolgus monkeys by subretinal injection reported dose-related anterior and posterior segment inflammation in animals, and cynomolgus monkeys were prematurely euthanized due to severe ocular inflammation [68]. In addition, vitreous aspirates from euthanized animals confirmed the presence of neutrophils and macrophages. Another study using a canine model similarly observed anterior and posterior uveitis following subretinal injection of AAV vector, with 3 of 17 eyes developing multifocal chorioretinitis, again associated with higher vector doses [69] ]. These findings strongly suggest that AAV vectors are under innate immune surveillance in the eye and can trigger deleterious inflammatory and immune responses.

Saline or similar amounts of ssAAV-eGFP or ss-AAV-eGFP-5x telomeres were delivered into neonatal eyes by subretinal injection and the expression of inflammatory and immune genes was measured. Three mice that received saline injections had consistently low Tnf expression in the retina and were set to 1-fold expression (FIG. 14A). In contrast, of the 5 mice that received ssAAV-eGFP, 2 exhibited mild Tnf upregulation (1.9-fold and 8.3-fold), while 3 animals exhibited large Tnf induction (62.2-fold, 534-fold and 1003-fold) , an average of 321 times for five animals. This finding suggests that while there are differences in inflammation, some animals developed very strong inflammatory responses in the retina following subretinal injection of ssAAV-eGFP. Variability may be attributed to differences in the immune status of each injection procedure or each animal. Surprisingly, the five animals that received ssAAV-eGFP-5x telomeres had an average Tnf induction rate of 5.6-fold with much less variability, suggesting that ssAAV-eGFP-5x telomeres were able to avoid triggering strong inflammation. Similar results were observed in the rest of the eye cup, but with lower magnitude, indicating that the inflammation was not limited to the retina (Figure 14B). We also measured expression in the retina of Ifng, a type II interferon critical for antiviral immune responses [70], and observed a similar pattern (Fig. 14C). Previous studies have shown that subretinal injection of AAV can trigger immune cell infiltration in the eye. Therefore, we also analyzed the expression of genes known to be specifically expressed in different types of immune cells. We found that the expression of Aif1 (encoding Iba1, a specific marker for microglia [71,72]) was up to 18.2-fold higher in ssAAV-eGFP injections compared to saline, suggesting that microglial cells play a role in the retina. Proliferation and/or activation (FIG. 15A). In contrast, ssAAV-eGFP-5x telomeres showed only a 1.9-fold induction of Aif1 expression. Additionally, we found that ssAAV-eGFP induced 45.8-fold and 41.8-fold induction of Cd4 and Cd8a (markers for CD4+ helper T cells and CD8+ cytolytic T cells, respectively), whereas ssAAV-eGFP-5x telomeres showed only 1.5-fold and 41.8-fold 3.4-fold induction (Figures 15B and 15C). Likewise, there were considerable differences between mice treated with ssAAV-eGFP, with a subgroup of animals showing strong induction. These results demonstrate that subretinal administration of ssAAV-eGFP stimulates T cell infiltration in the retina, which is strongly reduced by ssAAV-eGFP-5x telomeres. Taken together, our data demonstrate that while ssAAV-eGFP induces robust inflammatory and immune responses in the retina and surrounding tissues, and significant variability is observed, ssAAV-eGFP-5x telomeres are able to attenuate most of these responses.

Given the marked differences in inflammation in vitro and in vivo, we sought to determine whether there were any differences in long-term gene expression. 29 days after subretinal injection of mice, we examined eye cups plated on glass slides at P30 and found that more cells were GFP+ in ssAAV-eGFP-5x telomere-treated eyes compared to ssAAV-eGFP, And GFP expression was stronger, indicating enhanced gene expression (Figure 16). Thus, the engineered vectors were able to reduce inflammation and immune responses in the retina, and also enhance transgene expression.

Example 9 - Materials and Methods

animal

C57BL/6 mice (male, 7-9 weeks old) were purchased from Jackson Laboratory and CD1 mice were purchased from Charles River Laboratories.

AAV vector

Self-complementary (sc) or single-stranded (ss) AAV vectors were used in this study. Self-complementary vectors lack end-resolving sequences in one ITR. All vector genomes are flanked by AAV2 ITRs. scAAV-eGFP was purchased from Cell Biolabs (VPK-430) and has been described previously [73]. scAAV-eGFP expresses enhanced green fluorescent protein (eGFP) from the cytomegalovirus (CMV) promoter and includes the SV40 intron and SV40 polyA sequences. ssAAV-eGFP has been described previously [62] and was originally obtained from Harvard DF/HCC DNA Resource Core (clone ID: EvNO00061595). ssAAV-eGFP contains the CMV enhancer/promoter, human β-globin intron, eGFP and β-globin polyA sequences. "c41" (5'-TGGCGCGCACCCACGGCCTG-3'; SEQ ID NO: 1) from Pseudomonas aeruginosa and "telomeres" (5'-TTTAGGGTTAGGGTTAGGGTTAGGG-3') from mammalian telomeres have been described; SEQ ID NO: 9; the sequence of the initial T nucleotide is optional for function) [52,57,58,61]. A widely used "telomere" oligonucleotide (manufactured by Invivogen, catalog code "tlrl-ttag") has an additional T (bold) compared to published studies and is therefore included in the sequence. During the course of this study, Invivogen removed this extra T in its "telomere" oligonucleotide (catalog code "tlrl-ttag151"). Additionally, a "control" (5'-GCTAGATGTTAGCGT-3'; SEQ ID NO: 34) was used as a negative control sequence (Invivogen, cat. no. "tlr1-2088c") that did not inhibit TLR9 activation.

To design scAAV-eGFP, the sequence was inserted into a unique SpeI site found immediately 5' of the right ITR. To facilitate subcloning, a unique ClaI site was created immediately 5' of the insert, allowing ClaI/SpeI subcloning of the sequence. Three copies of "c41", "telomere" or "control" were inserted and separated with AAAAA linkers to obtain scAAV-eGFP-3xc41, scAAV-eGFP-3x telomere and scAAV-eGFP-3x control, respectively. Alternatively, insert a "telomere" copy with an AAAAA linker (SEQ ID NO: 8), resulting in scAAV-eGFP-1x telomeres. We also inserted a 3x telomere between the left ITR and the CMV promoter using a unique AvrII site, resulting in scAAV-3x telomere-eGFP.

To design ssAAV-eGFP, KpnI-5xtelomere(sense)-5xtelomere(antisense)-NheI was inserted immediately 5' of the XbaI site adjacent to the right ITR. Likewise, AAAAA is used as a linker between the "telomere" copies. Both sense and antisense sequences of "telomeres" are added, since single-stranded AAV vectors have an equal opportunity to package either the plus or minus strand of the viral genome, ensuring that all packaged AAV genomes will carry 5 in the correct orientation Copies of "telomeres".

The self-complementary vector was packaged into AAV2 (Vigene Biosciences) by triple transfection of HEK293 cells and purified using iodixanol gradient ultracentrifugation, then concentrated to 500 μl using an Amicon Ultra-15 column in PBS. Purified virus was titrated by qPCR using primers derived from ITR and AAV standards. The final yield of virus ranged from 0.5-3x10 ¹³ vg.

Single-stranded vectors were packaged into AAV8 based on previously described protocols [74,75]. Briefly, the AAV vector, rep2-cap8 packaging plasmid and adenovirus helper plasmid were transfected into HEK293T cells with polyethyleneimine, and the supernatant was collected 72 hours after transfection. AAV8 virus was precipitated with 8.5% w/v PEG8000 and 0.4M NaCl and centrifuged at 7000g. The pellet was resuspended in lysis buffer (150 mM NaCl and 20 mM Tris, pH 8.0) and MgCl2 was added to a final concentration of 1 mM. The resuspended virus was incubated with 25U/ml Benzonase (Sigma) for 15 minutes at 37°C and run on an iodixanol gradient. The recovered AAV vector was washed 3 times with PBS using an Amicon 100K column (EMD Millipore) and concentrated to 100-500 μl of PBS. For comparison, protein gels were performed to determine virus titers using serial dilutions of previous AAV standards.

Primary human monocytes and monocyte-derived macrophages for in vitro studies

Human peripheral blood mononuclear cells (PBMC) from unidentified healthy donors were purchased (ZenBio). The study was conducted in accordance with the ethical guidelines of Harvard Medical School. CD14+ monocytes were positively selected from PBMCs using anti-CD14 magnetic microbeads according to the manufacturer's instructions (Miltenyi Biotec), or CD14+ monocytes were purchased from Stemcell Technologies. To obtain monocyte-derived macrophages, monocytes were cultured with 50 ng/ml recombinant human macrophage colony stimulating factor (rhM-CSF, purchased from Peprotech) for 5-6 days to allow differentiation into macrophages. Monocytes and macrophages were used fresh or cryopreserved for subsequent studies.

1x105 monocytes or macrophages ^were seeded in 190 ul of RPMI growth medium per well in 96-well round bottom or 96-well flat bottom, respectively, and infected with 10 ul of AAV2 virus in PBS at the indicated MOI. Mock infection (10 μl of PBS added) and ODN2006 (5 μM final concentration, Invivogen), a CpG-containing oligonucleotide known to activate TLR9 and trigger inflammation, were used as negative and positive controls. Eighteen hours after infection, supernatants were collected and clarified by low speed centrifugation, followed by ELISA to detect human TNF (Thermo Scientific).

HeLa cell infection

HeLa cells are highly permissive for AAV2 vectors and are often used to determine transduction titers of AAV2 vector preparations [76]. Briefly, HeLa cells were seeded in 12 wells overnight and were approximately 80% confluent at the time of infection (~~ ^3x105 cells). Cells were infected with serial 10-fold dilutions of virus at the indicated MOIs and incubated for 48 h, then fixed with 1% paraformaldehyde in PBS before flow cytometric analysis of GFP+ cells. PBS mock-infected cells were used to determine GFP+ signal.

In vivo liver studies

Adult C57BL/6 mice were injected intravenously with 100 ul of PBS or AAV2 virus (10 ¹¹ vg/animal) via tail vein injection as previously described [36]. After 2 hours, animals were sacrificed and a portion of the right middle lobe of the liver was preserved in RNAlater solution (Thermo Scientific). Total RNA was extracted from 10-30 mg mechanically disrupted liver samples by using an RNA extraction kit (OMEGA Bio-Tek). Similar amounts of RNA were reverse transcribed into cDNA using a high-capacity RNA-to-cDNA kit (Thermo Scientific) and prepared using TaqMan Fast Advanced Master Mix (Thermo Scientific) and commercially available cDNAs for the indicated target genes (IDT). Pre-designed primers/probes with FAM reporter dyes were used to measure similar amounts of cDNA using quantitative PCR (qPCR). The expression level of each gene was calculated by normalizing against the housekeeping genes Actb or Gapdh using the ΔΔCT method and expressed as a fold level compared to saline-injected mice. All qPCR reactions were performed on a realplex ⁴ Mastercycle (Eppendorf).

In vivo eye studies

Subretinal injections were performed on postnatal day 1 (P1) CD1 neonatal eyes as described previously [74,75]. About 0.2 μl of AAV8 virus (1.8× ^{10 8} vg/eye) was introduced into the subretinal space using a glass pipette with a pull-out angle controlled by a FemtoJet (Eppendorf). At P21, animals were sacrificed and eye cups were dissected. RNA extraction, reverse transcription and qPCR were performed on the rest of the retina and eye cup as described in the liver study. To visualize GFP expression by histology, eyes were excised at P30, fixed in 4% paraformaldehyde for 2 h, and washed 3 times in PBS. The eye cup is dissected by removing the cornea, lens, iris, vitreous, and peripheral muscles. Images of eye cups flattened on glass slides were taken on a Keyence BZ-x700 microscope using an x10 objective. Images for between-group comparisons were taken under the same imaging settings in the same imaging group.

statistics

An unpaired two-tailed Student's t-test was used to compare the differences between the two unpaired experimental groups in all cases. A P value < 0.05 was considered statistically significant. No pre-specified effect size was assumed, and typically 3 to 5 replicates were used per condition.

This application relates to the following embodiments:

1. A nucleic acid molecule comprising:

A viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation.

2. A recombinant virus for delivering a desired function to mammalian cells, comprising:

A viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation.

3. The recombinant virus of embodiment 2, wherein the viral genome is an adeno-associated virus (AAV) genome.

4. The recombinant virus of embodiment 2, wherein the viral genome is selected from the group consisting of adenovirus, herpes simplex virus, varicella, variola virus, hepatitis B virus, cytomegalovirus, JC polyoma virus, BK polyoma virus, monkeypox virus , herpes zoster virus, Epstein-Barr virus, human herpesvirus 7, Kaposi's sarcoma-associated herpesvirus, and human parvovirus B19.

5. The recombinant virus of embodiment 2, wherein the viral genome is single-stranded.

6. The recombinant virus of embodiment 2, wherein the viral genome is packaged in a virion.

7. The recombinant virus of embodiment 2, wherein the viral genome comprises genes expressible in human cells.

8. The recombinant virus of embodiment 2, wherein the viral genome is a cytotoxic virus for lysing target tumor cells.

9. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises the c41 oligonucleotide sequence TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1).

10. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises multiple copies of the c41 sequence (SEQ ID NO: 1).

11. The recombinant virus of embodiment 10, wherein the inhibitory nucleic acid sequence comprises two copies of the c41 sequence (SEQ ID NO: 1) separated by a linker sequence.

12. The recombinant virus of embodiment 11, wherein the linker sequence is AAAAA (SEQ ID NO:8).

13. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is selected from the group consisting of:

ODN 2088: TCC TGG CGG GGA AGT (SEQ ID NO: 2);

ODN 4084-F: CCTGGATGGGAA (SEQ ID NO: 3);

ODN INH-1: CCTGGATGGGAATTCCCATCCAGG (SEQ ID NO: 4);

ODN INH-18: CCT GGA TGG GAA CTT ACC GCT GCA (SEQ ID NO: 5);

ODN TTAGGG:TT AGG GTT AGG GTT AGG GTT AGG G (SEQ ID NO: 6); and

G-ODN: CTC CTA TTG GGG GTT TCC TAT (SEQ ID NO: 7).

14. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is a bacterial sequence.

15. The recombinant virus of embodiment 2, wherein the viral genome comprises a non-human gene.

16. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is inserted downstream of or in the 3' untranslated region of the viral genome.

17. The recombinant virus of embodiment 2, wherein the viral genome is covalently linked to the inhibitory nucleic acid sequence through a phosphodiester bond.

18. The recombinant virus of embodiment 2, further comprising a detectable label.

19. The recombinant virus of embodiment 18, wherein the detectable marker is inducible.

20. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises the human telomere sequence set forth in SEQ ID NO:9.

21. The recombinant virus of embodiment 2, wherein the viral genome is self-complementary.

22. The recombinant virus of embodiment 2, wherein the viral genome is covalently linked to a plurality of inhibitory nucleic acid sequences.

23. The recombinant virus of embodiment 22, wherein the plurality of inhibitory nucleic acid sequences comprise inhibitory sequences and their reverse complements.

24. The recombinant virus of embodiment 10, wherein the inhibitory nucleic acid sequence comprises 3 copies of the c41 sequence (SEQ ID NO: 1), each copy being separated by a linker sequence.

25. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is selected from the group consisting of:

ODN 2114: TCCTGGAGGGGAAGT (SEQ ID NO: 16);

ODN 4024: TCCTGGATGGGAAGT (SEQ ID NO: 17);

ODN INH-4: TTCCCATCCAGGCCTGGATGGGAA (SEQ ID NO: 18);

ODN INH-13: CTTACCGCTGCACCTGGATGGGAA (SEQ ID NO: 19);

ODN Poly-G: GGGGGGGGGGGGGGGGGGGG (SEQ ID NO: 20);

ODN GpG: TGACTGTGAAGGTTAGAGATGA (SEQ ID NO: 21);

ODN IRS-869: TCCTGGAGGGGTTGT (SEQ ID NO: 22);

ODN IRS-954:TGCTCCTGGAGGGGTTGT (SEQ ID NO:23); and

ODN 21158: CCTGGCGGGG (SEQ ID NO: 24).

26. The recombinant virus of embodiment 13, wherein the inhibitory nucleic acid sequence is ODN TTAGGG (SEQ ID NO: 6).

27. The recombinant virus of embodiment 26, wherein the inhibitory sequence is covalently linked to a linker.

28. The recombinant virus of embodiment 26, wherein the inhibitory sequence is upstream of the linker.

29. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises multiple copies of ODN TTAGGG (SEQ ID NO: 6).

30. The recombinant virus of embodiment 29, wherein the multiple copies of ODN TTAGGG (SEQ ID NO: 6) are each separated by a linker.

31. The recombinant virus of embodiment 30, wherein the inhibitory nucleic acid sequence comprises at least 2, at least 3, at least 4 or at least 5 copies of ODN TTAGGG (SEQ ID NO: 6), each copy being separated by a linker .

32. The recombinant virus of embodiment 30, wherein the linker is AAAAA.

33. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is a human sequence.

34. The recombinant virus of embodiment 2, wherein the viral genome comprises a non-human nucleic acid sequence.

35. The recombinant virus of embodiment 2, wherein the viral genome comprises human genes.

36. The recombinant virus of embodiment 2, wherein the viral genome comprises human nucleic acid sequences.

37. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is inserted in the 5' untranslated region of the viral genome.

38. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is inserted upstream of a promoter in the viral genome.

39. The recombinant virus of embodiment 2, further comprising an inducible promoter.

40. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises two repeat monomers of SEQ ID NO:1.

41. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises the three repeat monomers of SEQ ID NO:1.

42. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises SEQ ID NO:6 or SEQ ID NO:9.

43. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises the three repeat monomers of SEQ ID NO:6 or SEQ ID NO:9.

44. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence comprises five repeat monomers of SEQ ID NO:6 or SEQ ID NO:9.

45. A method of treating a mammal comprising:

The recombinant virus of Embodiment 2 is administered to a mammal in need thereof.

46. The method of embodiment 45, wherein the viral genome is an adeno-associated virus (AAV) genome.

47. The method of embodiment 45, wherein the viral genome is selected from the group consisting of adenovirus, herpes simplex virus, varicella, variola virus, hepatitis B virus, cytomegalovirus, JC polyoma virus, BK polyoma virus, monkeypox virus, zonavirus Herpes virus, Epstein-Barr virus, human herpes virus 7, Kaposi's sarcoma-associated herpes virus, and human parvovirus B19.

48. The method of embodiment 45, wherein the viral genome is single-stranded.

49. The method of embodiment 45, wherein the viral genome is packaged in a virion.

50. The method of embodiment 45, wherein the viral genome is a gene therapy vector for delivering expressible genes to human cells.

51. The method of embodiment 45, wherein the viral genome is a cytotoxic virus for lysing target tumor cells.

52. The method of embodiment 45, wherein the inhibitory nucleic acid sequence comprises a c41 oligonucleotide sequence (SEQ ID NO: 1).

53. The method of embodiment 45, wherein the inhibitory nucleic acid sequence comprises multiple copies of the c41 sequence (SEQ ID NO: 1).

54. The method of embodiment 45, wherein the inhibitory nucleic acid sequence comprises two copies of the c41 sequence (SEQ ID NO: 1) separated by a linker sequence.

55. The method of embodiment 54, wherein the linker sequence is AAAAA (SEQ ID NO:8).

56. The method of embodiment 45, wherein the inhibitory nucleic acid sequence is a bacterial sequence.

57. The method of embodiment 45, wherein the viral genome comprises genes for delivery to and expression in human cells.

58. The method of embodiment 45, wherein the administering step is repeated.

59. A method of preparing a viral genome of a recombinant virus, comprising:

An inhibitory nucleic acid sequence is inserted into the viral genome, wherein the inhibitory nucleic acid sequence binds to TLR9 but does not trigger TLR9 activation.

60. The method of embodiment 59, further comprising:

The viral genome is packaged in virions.

61. The method of embodiment 59, wherein the inserting step uses DNA ligase.

62. The method of embodiment 59, wherein the viral genome is selected from the group consisting of adenovirus, herpes simplex virus, varicella, smallpox virus, hepatitis B virus, cytomegalovirus, JC polyoma virus, BK polyoma virus, monkeypox virus, Herpes zoster virus, Epstein-Barr virus, human herpesvirus 7, Kaposi's sarcoma-associated herpesvirus, and human parvovirus B19.

63. The method of embodiment 59, wherein the viral genome is single-stranded when in a virion.

64. The method of embodiment 59, wherein the viral genome is a gene therapy vector for delivering expressible genes to human cells.

65. The method of embodiment 59, wherein the viral genome is a cytotoxic virus for lysing target tumor cells.

66. The method of embodiment 59, wherein the inhibitory nucleic acid sequence comprises a c41 oligonucleotide sequence (SEQ ID NO: 1).

67. The method of embodiment 59, wherein the inhibitory nucleic acid sequence comprises multiple copies of the c41 sequence (SEQ ID NO: 1).

68. The method of embodiment 59, wherein the inhibitory nucleic acid sequence comprises two copies of the c41 sequence (SEQ ID NO: 1) separated by a linker sequence.

69. The method of embodiment 68, wherein the linker sequence is AAAAA (SEQ ID NO:8).

70. The method of embodiment 59, wherein the inhibitory nucleic acid sequence is a bacterial sequence.

71. The method of embodiment 59, wherein the viral genome of the recombinant virus comprises genes for delivery to and expression in human cells.

72. A nucleic acid molecule comprising:

Inverted terminal repeats (ITRs) and nucleic acid sequences that inhibit TLR9-mediated inflammation.

73. A recombinant virus for delivering a desired function to a mammalian cell, comprising:

Viral genomes comprising nucleic acid sequences that inhibit TLR9-mediated inflammation.

74. A method of preparing a viral genome of a recombinant virus, comprising:

A nucleic acid sequence is inserted into the viral genome, wherein the nucleic acid sequence inhibits TLR9-mediated inflammation.

75. A nucleic acid vector comprising at least one nucleic acid sequence capable of inhibiting TLR9-mediated inflammation.

76. The nucleic acid vector of embodiment 75, comprising a plurality of said nucleic acid sequences.

77. The nucleic acid vector of embodiment 75, wherein a linker separates each of said nucleic acid sequences.

78. The nucleic acid vector of embodiment 75, comprising at least 2, at least 3, at least 4, or at least 5 copies of the nucleic acid sequence.

79. The nucleic acid vector of embodiment 75, wherein the nucleic acid is covalently linked to the gene.

80. A method of reducing the immunogenicity of a modified virus having a genome, the method comprising inserting a nucleic acid sequence in the genome, the inhibitory nucleic acid sequence inhibits TLR9-mediated inflammation, whereby the The modified virus elicits a reduced inflammatory response in the host compared to a virus that does not contain the nucleic acid sequence.

81. A method of increasing the expression of a transgene introduced by a virus in a host cell, the method comprising introducing into the host a modified virus with a genome comprising a nucleic acid sequence that inhibits TLR9 mediation inflammation, whereby the modified virus results in higher expression of the transgene in the host cell compared to a virus that does not contain the nucleic acid sequence.

82. A composition comprising a viral capsid encapsulating a nucleic acid sequence that inhibits TLR9-mediated inflammation.

83. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is 95% identical to the c41 oligonucleotide sequence TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1).

84. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is 95% identical to SEQ ID NO:9.

85. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is 95% identical to a sequence selected from the group consisting of:

ODN 2088: TCC TGG CGG GGA AGT (SEQ ID NO: 2);

ODN 4084-F: CCTGGATGGGAA (SEQ ID NO: 3);

ODN INH-1: CCTGGATGGGAATTCCCATCCAGG (SEQ ID NO: 4);

ODN INH-18: CCT GGA TGG GAA CTT ACC GCT GCA (SEQ ID NO: 5);

ODN TTAGGG:TT AGG GTT AGG GTT AGG GTT AGG G (SEQ ID NO: 6); and

G-ODN: CTC CTA TTG GGG GTT TCC TAT (SEQ ID NO: 7).

86. The recombinant virus of embodiment 2, wherein the inhibitory nucleic acid sequence is 95% identical to a sequence selected from the group consisting of:

ODN 2114: TCCTGGAGGGGAAGT (SEQ ID NO: 16);

ODN 4024: TCCTGGATGGGAAGT (SEQ ID NO: 17);

ODN INH-4: TTCCCATCCAGGCCTGGATGGGAA (SEQ ID NO: 18);

ODN INH-13: CTTACCGCTGCACCTGGATGGGAA (SEQ ID NO: 19);

ODN Poly-G: GGGGGGGGGGGGGGGGGGGG (SEQ ID NO: 20);

ODN GpG: TGACTGTGAAGGTTAGAGATGA (SEQ ID NO: 21);

ODN IRS-869: TCCTGGAGGGGTTGT (SEQ ID NO: 22);

ODN IRS-954:TGCTCCTGGAGGGGTTGT (SEQ ID NO:23); and

ODN 21158: CCTGGCGGGG (SEQ ID NO: 24).

87. The method of embodiment 45, wherein the inhibitory nucleic acid sequence comprises SEQ ID NO:6 or SEQ ID NO:9.

88. The method of embodiment 45, wherein the inhibitory nucleic acid sequence comprises multiple copies of SEQ ID NO:6 and/or SEQ ID NO:9.

89. The method of embodiment 45, wherein the inhibitory nucleic acid sequence comprises two copies of SEQ ID NO:6 and/or SEQ ID NO:9 separated by a linker sequence.

90. The method of embodiment 45, wherein the inhibitory nucleic acid sequence is a human sequence.

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68. Ye GJ, Budzynski E, Sonnentag P, Nork TM, Miller PE, Sharma AK, etal. Safety and Biodistribution Evaluation in Cynomolgus Macaques of rAAV2tYF-PR1.7-hCNGB3, a Recombinant AAV Vector for Treatment of Achromatopsia. Hum GeneTher Clin Dev.2016.doi:10.1089/hum.2015.164.PubMed PMID:26956923.

69. Komaromy AM, Alexander JJ, Rowlan JS, Garcia MM, Chiodo VA, Kaya A, et al. Gene therapy rescues cone function in congenital achromatopsia. Hum MolGenet. 2010;19(13):2581-93.doi:10.1093/hmg /ddq136.PubMed PMID:20378608;PubMedCentral PMCID:PMCPMC2883338.

70. Schoenborn JR, Wilson CB. Regulation of interferon-gamma during innate and adaptive immune responses. Adv Immunol. 2007;96:41-101.doi:10.1016/S0065-2776(07)96002-2.PubMed PMID:17981204.

71. Imai Y, Ibata I, Ito D, Ohsawa K, Kohsaka S. A novel gene iba1 in themajor histocompatibility complex class III region encoding an EF hand protein expressed in a monocytic lineage. Biochem Biophys Res Commun. 1996;224(3):855- 62.doi:10.1006/bbrc.1996.1112.PubMed PMID:8713135.

72. Ito D, Imai Y, Ohsawa K, Nakajima K, Fukuuchi Y, Kohsaka S. Microglia-specific localisation of a novel calcium binding protein, Iba1. Brain Res MolBrain Res. 1998;57(1):1-9.PubMed PMID: 9630473.

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sequence listing

<110> Chan, Ying Kai

Church, George

<120> Engineered viral vectors reduce the induction of inflammation and immune responses

<130> 010498.00962

<160> 34

<170> FastSEQ for Windows Version 4.0

<210> 1

<211> 20

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 1

tggcgcgcac ccacggcctg 20

<210> 2

<211> 15

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 2

tcctggcggg gaagt 15

<210> 3

<211> 12

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 3

cctggatggg aa 12

<210> 4

<211> 24

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 4

cctggatggg aattcccatc cagg 24

<210> 5

<211> 24

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 5

cctggatggg aacttaccgc tgca 24

<210> 6

<211> 24

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 6

ttagggttag ggttagggtt aggg 24

<210> 7

<211> 21

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 7

ctcctattgg gggtttccta t 21

<210> 8

<211> 5

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 8

aaaaa 5

<210> 9

<211> 25

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 9

tttagggtta gggttagggt taggg 25

<210> 10

<211> 4369

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 10

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaactagt ccactccctc tctgcgcgct cgctcgctca ctgaggccgg 1620

gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc 1680

gcgcagagag ggacagatcc gggcccgcat gcgtcgacaa ttcactggcc gtcgttttac 1740

aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc 1800

ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc 1860

gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta 1920

tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc 1980

agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat 2040

ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt 2100

catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg 2160

tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa 2220

cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac 2280

cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg 2340

tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc 2400

tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg 2460

atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga 2520

gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc 2580

aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag 2640

aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga 2700

gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg 2760

cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga 2820

atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt 2880

tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact 2940

ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt 3000

ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg 3060

ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta 3120

tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac 3180

tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta 3240

aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt 3300

tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt 3360

tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt 3420

gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc 3480

agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg 3540

tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg 3600

ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt 3660

cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac 3720

tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg 3780

acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg 3840

gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat 3900

ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt 3960

tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg 4020

attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa 4080

cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc 4140

ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga 4200

aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg 4260

ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc 4320

acacaggaaa cagctatgac catgattacg ccaagctctc gagatctag 4369

<210> 11

<211> 4450

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 11

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat tggcgcgcac ccacggcctg aaaaatggcg cgcacccacg 1620

gcctgaaaaa tggcgcgcac ccacggcctg aaaaaactag tccactccct ctctgcgcgc 1680

tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc 1740

ggcctcagtg agcgagcgag cgcgcagaga gggacagatc cgggcccgca tgcgtcgaca 1800

attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta 1860

atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg 1920

atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgcctgatg cggtattttc 1980

tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct 2040

ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac 2100

gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca 2160

tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac 2220

gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt 2280

ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt 2340

atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta 2400

tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg 2460

ttttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac 2520

gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg 2580

aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc 2640

gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg 2700

ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat 2760

gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg 2820

gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg 2880

atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc 2940

ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt 3000

cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct 3060

cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc 3120

gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca 3180

cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct 3240

cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt 3300

taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga 3360

ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca 3420

aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac 3480

caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg 3540

taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag 3600

gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac 3660

cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt 3720

taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg 3780

agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc 3840

ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc 3900

gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc 3960

acctctgact tgagcgtcga ttttttgtgat gctcgtcagg ggggcggagc ctatggaaaa 4020

acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt 4080

tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg 4140

ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag 4200

agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc 4260

acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc 4320

tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa 4380

ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct 4440

cgagatctag 4450

<210> 12

<211> 4465

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 12

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat tttagggtta gggttagggt tagggaaaaa tttagggtta 1620

gggttagggt tagggaaaaa tttagggtta gggttagggt tagggaaaaa actagtccac 1680

tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc 1740

gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agagagggac agatccgggc 1800

ccgcatgcgt cgacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg 1860

gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg 1920

aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc 1980

tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc 2040

tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg 2100

ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg 2160

tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa 2220

agggcctcgt gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga 2280

cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa 2340

tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt 2400

gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg 2460

cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag 2520

atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg 2580

agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg 2640

gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt 2700

ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga 2760

cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac 2820

ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc 2880

atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc 2940

gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac 3000

tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag 3060

gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg 3120

gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta 3180

tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg 3240

ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata 3300

tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt 3360

ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc 3420

ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct 3480

tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa 3540

ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag 3600

tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc 3660

tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg 3720

actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca 3780

cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat 3840

gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg 3900

tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc 3960

ctgtcgggtt tcgccacctc tgacttgagc gtcgatttttt gtgatgctcg tcaggggggc 4020

ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc 4080

cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg 4140

cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga 4200

gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc 4260

attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa 4320

ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc 4380

gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg 4440

attacgccaa gctctcgaga tctag 4465

<210> 13

<211> 4465

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 13

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggtt tagggttagg gttagggtta gggaaaaatt tagggttagg gttagggtta 240

gggaaaaatt tagggttagg gttagggtta gggaaaaacc taggcatatg ccaagtacgc 300

cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct 360

tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga 420

tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa 480

gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc 540

caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg 600

aggtctatat aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccgg 660

actctaaggt aaatataaaa ttttttaagtg tataatgtgt taaactactg attctaattg 720

tttctctctt ttagattcca acctttggaa ctgaattccg cgggcccggg atccaccggt 780

cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga 840

gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc 900

cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg 960

gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca 1020

catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac 1080

catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga 1140

caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct 1200

ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca 1260

gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca 1320

gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga 1380

caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca 1440

catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta 1500

caagtaaagc ggccgctagg cctcacctgc gatctcgatg ctttatttgt gaaatttgtg 1560

atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt 1620

gcattcattt tatgtttcag gttcagggggg aggtgtggga ggttttttaa actagtccac 1680

tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc 1740

gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agagagggac agatccgggc 1800

ccgcatgcgt cgacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg 1860

gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg 1920

aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc 1980

tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc 2040

tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg 2100

ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg 2160

tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa 2220

agggcctcgt gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga 2280

cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa 2340

tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt 2400

gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg 2460

cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag 2520

atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg 2580

agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg 2640

gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt 2700

ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga 2760

cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac 2820

ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc 2880

atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc 2940

gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac 3000

tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag 3060

gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg 3120

gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta 3180

tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg 3240

ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata 3300

tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt 3360

ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc 3420

ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct 3480

tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa 3540

ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag 3600

tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc 3660

tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg 3720

actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca 3780

cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat 3840

gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg 3900

tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc 3960

ctgtcgggtt tcgccacctc tgacttgagc gtcgatttttt gtgatgctcg tcaggggggc 4020

ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc 4080

cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg 4140

cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga 4200

gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc 4260

attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa 4320

ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc 4380

gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg 4440

attacgccaa gctctcgaga tctag 4465

<210> 14

<211> 6940

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 14

cagcagctgc gcgctcgctc gctcactgag gccgccccggg caaagcccgg gcgtcgggcg 60

acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120

atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180

tgctctagcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat gagcttggcc 240

cattgcatac gttgtatcca tatcataata tgtacattta tattggctca tgtccaacat 300

taccgccatg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat 360

tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg 420

gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa 480

cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact 540

tggcagtaca tcaagtgtat catatgccaa gtacgcccccc tattgacgtc aatgacggta 600

aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt 660

acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg 720

ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg 780

ggagttttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc 840

cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt 900

tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac 960

accgggaccg atccagcctc ccctcgaagc tgatcctgag aacttcaggg tgagtctatg 1020

ggacccttga tgttttcttt ccccttcttt tctatggtta agttcatgtc ataggaaggg 1080

gagaagtaac agggtacaca tattgaccaa atcagggtaa ttttgcattt gtaattttaa 1140

aaaatgcttt cttcttttaa tatacttttt tgtttatctt atttctaata ctttccctaa 1200

tctctttctt tcagggcaat aatgatacaa tgtatcatgc ctctttgcac cattctaaag 1260

aataacagtg ataatttctg ggttaaggca atagcaatat ttctgcatat aaatatttct 1320

gcatataaat tgtaactgat gtaagaggtt tcatattgct aatagcagct acaatccagc 1380

taccattctg cttttatttt atggttggga taaggctgga ttattctgag tccaagctag 1440

gcccttttgc taatcatgtt catacctctt atcttcctcc cacagctcct gggcaacgtg 1500

ctggtctgtg tgctggccca tcactttggc aaagaattcc gcgggcccgg gatccaccgg 1560

tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 1620

agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 1680

ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 1740

ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 1800

acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 1860

ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 1920

acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 1980

tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 2040

agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 2100

agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 2160

acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 2220

acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 2280

acaagtaaag cggccgctct agaggatcca agcttatcga taccgtcgac ctcgagggcc 2340

cagatctaat tcaccccacc agtgcaggct gcctatcaga aagtggtggc tggtgtggct 2400

aatgccctgg cccacaagta tcactaagct cgctttcttg ctgtccaatt tctattaaag 2460

gttcctttgt tccctaagtc caactactaa actgggggat attatgaagg gccttgagca 2520

tctggattct gcctaataaa aaacatttat tttcattgca atgatgtatt taaattattt 2580

ctgaatattt tactaaaaag ggaatgtggg aggtcagtgc atttaaaaca taaagaaatg 2640

aagagctagt tcaaaccttg ggaaaataca ctatatctta aactccatga aagaaggtga 2700

ggctgcaaac agctaatgca cattggcaac agcccctgat gcctatgcct tattcatccc 2760

tcagaaaagg attcaagtag aggcttgatt tggaggttaa agttttgcta tgctgtattt 2820

tacattactt attgttttag ctgtcctcat gaatgtcttt tcactaccca tttgcttatc 2880

ctgcatctct cagccttgac tccactcagt tctcttgctt agagatacca cctttcccct 2940

gaagtgttcc ttccatgttt tacggcgaga tggtttctcc tcgcctggcc actcagcctt 3000

agttgtctct gttgtcttat agaggtctac ttgaagaagg aaaaacaggg ggcatggttt 3060

gactgtcctg tgagcccttc ttccctgcct cccccactca cagtgacccg gaatccctcg 3120

acatctagag catggctacg tagataagta gcatggcggg ttaatcatta actacaagga 3180

acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg 3240

gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc 3300

gcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca 3360

gcctgaatgg cgaatggaat tccagacgat tgagcgtcaa aatgtaggta tttccatgag 3420

cgtttttcct gttgcaatgg ctggcggtaa tattgttctg gatattacca gcaaggccga 3480

tagtttgagt tcttctactc aggcaagtga tgttattact aatcaaagaa gtattgcgac 3540

aacggttaat ttgcgtgatg gacagactct tttactcggt ggcctcactg attataaaaa 3600

cacttctcag gattctggcg taccgttcct gtctaaaatc cctttaatcg gcctcctgtt 3660

tagctcccgc tctgattcta acgaggaaag cacgttatac gtgctcgtca aagcaaccat 3720

agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 3780

ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 3840

ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 3900

ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg 3960

ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata 4020

gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt 4080

tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat 4140

ttaacgcgaa ttttaacaaa atattaacgt ttacaattta aatatttgct tatacaatct 4200

tcctgttttt ggggcttttc tgattatcaa ccggggtaca tatgattgac atgctagttt 4260

tacgattacc gttcatcgat tctcttgttt gctccagact ctcaggcaat gacctgatag 4320

cctttgtaga gacctctcaa aaatagctac cctctccggc atgaatttat cagctagaac 4380

ggttgaatat catattgatg gtgatttgac tgtctccggc ctttctcacc cgtttgaatc 4440

tttacctaca cattactcag gcattgcatt taaaatatat gagggttcta aaaattttta 4500

tccttgcgtt gaaataaagg cttctcccgc aaaagtatta cagggtcata atgtttttgg 4560

tacaaccgat ttagctttat gctctgaggc tttattgctt aattttgcta attctttgcc 4620

ttgcctgtat gatttattgg atgttggaat tcctgatgcg gtattttctc cttacgcatc 4680

tgtgcggtat ttcacaccgc atatggtgca ctctcagtac aatctgctct gatgccgcat 4740

agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc 4800

tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt 4860

tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat 4920

aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg 4980

tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga 5040

gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac 5100

atttccgtgt cgcccttatt ccctttttttg cggcattttg ccttcctgtt tttgctcacc 5160

cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca 5220

tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc 5280

caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg 5340

ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac 5400

cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca 5460

taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg 5520

agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac 5580

cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg 5640

caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat 5700

taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg 5760

ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg 5820

cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc 5880

aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc 5940

attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt 6000

tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt 6060

aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt 6120

gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag 6180

cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca 6240

gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca 6300

agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg 6360

ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg 6420

cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct 6480

acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga 6540

gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc 6600

ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg 6660

agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg 6720

cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt 6780

tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc 6840

gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac 6900

gcaaaccgcc tctccccgcg cgttggccga ttcattaatg 6940

<210> 15

<211> 7257

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 15

cagcagctgc gcgctcgctc gctcactgag gccgccccggg caaagcccgg gcgtcgggcg 60

acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120

atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180

tgctctagcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat gagcttggcc 240

cattgcatac gttgtatcca tatcataata tgtacattta tattggctca tgtccaacat 300

taccgccatg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat 360

tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg 420

gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa 480

cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact 540

tggcagtaca tcaagtgtat catatgccaa gtacgcccccc tattgacgtc aatgacggta 600

aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt 660

acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg 720

ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg 780

ggagttttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc 840

cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt 900

tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac 960

accgggaccg atccagcctc ccctcgaagc tgatcctgag aacttcaggg tgagtctatg 1020

ggacccttga tgttttcttt ccccttcttt tctatggtta agttcatgtc ataggaaggg 1080

gagaagtaac agggtacaca tattgaccaa atcagggtaa ttttgcattt gtaattttaa 1140

aaaatgcttt cttcttttaa tatacttttt tgtttatctt atttctaata ctttccctaa 1200

tctctttctt tcagggcaat aatgatacaa tgtatcatgc ctctttgcac cattctaaag 1260

aataacagtg ataatttctg ggttaaggca atagcaatat ttctgcatat aaatatttct 1320

gcatataaat tgtaactgat gtaagaggtt tcatattgct aatagcagct acaatccagc 1380

taccattctg cttttatttt atggttggga taaggctgga ttattctgag tccaagctag 1440

gcccttttgc taatcatgtt catacctctt atcttcctcc cacagctcct gggcaacgtg 1500

ctggtctgtg tgctggccca tcactttggc aaagaattcc gcgggcccgg gatccaccgg 1560

tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 1620

agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 1680

ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 1740

ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 1800

acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 1860

ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 1920

acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 1980

tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 2040

agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 2100

agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 2160

acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 2220

acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 2280

acaagtaaag cggccgctct agaggatcca agcttatcga taccgtcgac ctcgagggcc 2340

cagatctaat tcaccccacc agtgcaggct gcctatcaga aagtggtggc tggtgtggct 2400

aatgccctgg cccacaagta tcactaagct cgctttcttg ctgtccaatt tctattaaag 2460

gttcctttgt tccctaagtc caactactaa actgggggat attatgaagg gccttgagca 2520

tctggattct gcctaataaa aaacatttat tttcattgca atgatgtatt taaattattt 2580

ctgaatattt tactaaaaag ggaatgtggg aggtcagtgc atttaaaaca taaagaaatg 2640

aagagctagt tcaaaccttg ggaaaataca ctatatctta aactccatga aagaaggtga 2700

ggctgcaaac agctaatgca cattggcaac agcccctgat gcctatgcct tattcatccc 2760

tcagaaaagg attcaagtag aggcttgatt tggaggttaa agttttgcta tgctgtattt 2820

tacattactt attgttttag ctgtcctcat gaatgtcttt tcactaccca tttgcttatc 2880

ctgcatctct cagccttgac tccactcagt tctcttgctt agagatacca cctttcccct 2940

gaagtgttcc ttccatgttt tacggcgaga tggtttctcc tcgcctggcc actcagcctt 3000

agttgtctct gttgtcttat agaggtctac ttgaagaagg aaaaacaggg ggcatggttt 3060

gactgtcctg tgagcccttc ttccctgcct cccccactca cagtgacccg gaatccctcg 3120

acaggtacct ttagggttag ggttagggtt agggaaaaat ttagggttag ggttagggtt 3180

agggaaaaat ttagggttag ggttagggtt agggaaaaat ttagggttag ggttagggtt 3240

agggaaaaat ttagggttag ggttagggtt agggaaaaac cctaacccta accctaaccc 3300

taaatttttt ccctaaccct aaccctaacc ctaaattttt tccctaaccc taaccctaac 3360

cctaaatttt ttccctaacc ctaaccctaa ccctaaattt tttccctaac cctaacccta 3420

accctaaatt ttttgctagc tctagagcat ggctacgtag ataagtagca tggcgggtta 3480

atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc 3540

tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc 3600

tcagtgagcg agcgagcgcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc 3660

ttcccaacag ttgcgcagcc tgaatggcga atggaattcc agacgattga gcgtcaaaat 3720

gtaggtattt ccatgagcgt ttttcctgtt gcaatggctg gcggtaatat tgttctggat 3780

attaccagca aggccgatag tttgagttct tctactcagg caagtgatgt tattactaat 3840

caaagaagta ttgcgacaac ggttaatttg cgtgatggac agactctttt actcggtggc 3900

ctcactgatt ataaaaacac ttctcaggat tctggcgtac cgttcctgtc taaaatccct 3960

ttaatcggcc tcctgtttag ctcccgctct gattctaacg aggaaagcac gttatacgtg 4020

ctcgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt 4080

ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt 4140

cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct 4200

ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg 4260

tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga 4320

gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc 4380

ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga 4440

gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caatttaaat 4500

atttgcttat acaatcttcc tgtttttggg gcttttctga ttatcaaccg gggtacatat 4560

gattgacatg ctagttttac gattaccgtt catcgattct cttgtttgct ccagactctc 4620

aggcaatgac ctgatagcct ttgtagagac ctctcaaaaa tagctaccct ctccggcatg 4680

aatttatcag ctagaacggt tgaatatcat attgatggtg atttgactgt ctccggcctt 4740

tctcacccgt ttgaatcttt acctacacat tactcaggca ttgcatttaa aatatatgag 4800

ggttctaaaa atttttatcc ttgcgttgaa ataaaggctt ctcccgcaaa agtattacag 4860

ggtcataatg tttttggtac aaccgattta gctttatgct ctgaggcttt attgcttaat 4920

tttgctaatt ctttgccttg cctgtatgat ttattggatg ttggaattcc tgatgcggta 4980

ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc tcagtacaat 5040

ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc 5100

ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag 5160

ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt 5220

gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg 5280

cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa 5340

tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa 5400

gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct 5460

tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg 5520

tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg 5580

ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt 5640

atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga 5700

cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga 5760

attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac 5820

gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg 5880

ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac 5940

gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct 6000

agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct 6060

gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg 6120

gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat 6180

ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg 6240

tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat 6300

tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct 6360

catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa 6420

gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa 6480

aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc 6540

gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta 6600

gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct 6660

gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg 6720

atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag 6780

cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc 6840

cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg 6900

agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt 6960

tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg 7020

gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca 7080

catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg 7140

agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc 7200

ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatg 7257

<210> 16

<211> 15

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 16

tcctggaggg gaagt 15

<210> 17

<211> 15

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 17

tcctggatgg gaagt 15

<210> 18

<211> 24

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 18

ttcccatcca ggcctggatg ggaa 24

<210> 19

<211> 24

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 19

cttaccgctg cacctggatg ggaa 24

<210> 20

<211> 20

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 20

gggggggggg gggggggggg 20

<210> 21

<211> 22

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 21

tgactgtgaa ggttagagat ga 22

<210> 22

<211> 15

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 22

tcctggaggg gttgt 15

<210> 23

<211> 18

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 23

tgctcctgga ggggttgt 18

<210> 24

<211> 10

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 24

cctggcgggg 10

<210> 25

<211> 4369

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 25

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaactagt ccactccctc tctgcgcgct cgctcgctca ctgaggccgg 1620

gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc 1680

gcgcagagag ggacagatcc gggcccgcat gcgtcgacaa ttcactggcc gtcgttttac 1740

aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc 1800

ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc 1860

gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta 1920

tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc 1980

agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat 2040

ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt 2100

catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg 2160

tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa 2220

cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac 2280

cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg 2340

tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc 2400

tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg 2460

atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga 2520

gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc 2580

aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag 2640

aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga 2700

gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg 2760

cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga 2820

atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt 2880

tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact 2940

ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt 3000

ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg 3060

ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta 3120

tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac 3180

tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta 3240

aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt 3300

tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt 3360

tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt 3420

gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc 3480

agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg 3540

tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg 3600

ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt 3660

cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac 3720

tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg 3780

acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg 3840

gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat 3900

ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt 3960

tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg 4020

attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa 4080

cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc 4140

ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga 4200

aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg 4260

ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc 4320

acacaggaaa cagctatgac catgattacg ccaagctctc gagatctag 4369

<210> 26

<211> 4450

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 26

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat tggcgcgcac ccacggcctg aaaaatggcg cgcacccacg 1620

gcctgaaaaa tggcgcgcac ccacggcctg aaaaaactag tccactccct ctctgcgcgc 1680

tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc 1740

ggcctcagtg agcgagcgag cgcgcagaga gggacagatc cgggcccgca tgcgtcgaca 1800

attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta 1860

atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg 1920

atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgcctgatg cggtattttc 1980

tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct 2040

ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac 2100

gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca 2160

tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac 2220

gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt 2280

ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt 2340

atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta 2400

tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg 2460

ttttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac 2520

gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg 2580

aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc 2640

gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg 2700

ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat 2760

gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg 2820

gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg 2880

atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc 2940

ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt 3000

cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct 3060

cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc 3120

gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca 3180

cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct 3240

cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt 3300

taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga 3360

ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca 3420

aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac 3480

caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg 3540

taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag 3600

gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac 3660

cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt 3720

taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg 3780

agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc 3840

ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc 3900

gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc 3960

acctctgact tgagcgtcga ttttttgtgat gctcgtcagg ggggcggagc ctatggaaaa 4020

acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt 4080

tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg 4140

ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag 4200

agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc 4260

acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc 4320

tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa 4380

ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct 4440

cgagatctag 4450

<210> 27

<211> 4425

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 27

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat tggcgcgcac ccacggcctg aaaaatggcg cgcacccacg 1620

gcctgaaaaa actagtccac tccctctctg cgcgctcgct cgctcactga ggccgggcga 1680

ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc 1740

agagagggac agatccgggc ccgcatgcgt cgacaattca ctggccgtcg ttttacaacg 1800

tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt 1860

cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag 1920

cctgaatggc gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc 1980

acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc 2040

ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc 2100

ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc 2160

accgaaacgc gcgagacgaa agggcctcgt gatacgccta ttttttatagg ttaatgtcat 2220

gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc 2280

tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg 2340

ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc 2400

ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt 2460

gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct 2520

caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac 2580

ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact 2640

cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa 2700

gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga 2760

taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt 2820

tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga 2880

agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg 2940

caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat 3000

ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat 3060

tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc 3120

agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga 3180

tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc 3240

agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 3300

gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc 3360

gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt 3420

tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt 3480

gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 3540

accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 3600

accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 3660

gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 3720

ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 3780

atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 3840

gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 3900

cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 3960

gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 4020

gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 4080

tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac 4140

cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct 4200

ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc 4260

gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt 4320

acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac 4380

aggaaacagc tatgaccatg attacgccaa gctctcgaga tctag 4425

<210> 28

<211> 4465

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 28

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat tttagggtta gggttagggt tagggaaaaa tttagggtta 1620

gggttagggt tagggaaaaa tttagggtta gggttagggt tagggaaaaa actagtccac 1680

tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc 1740

gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agagagggac agatccgggc 1800

ccgcatgcgt cgacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg 1860

gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg 1920

aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc 1980

tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc 2040

tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg 2100

ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg 2160

tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa 2220

agggcctcgt gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga 2280

cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa 2340

tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt 2400

gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg 2460

cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag 2520

atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg 2580

agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg 2640

gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt 2700

ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga 2760

cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac 2820

ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc 2880

atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc 2940

gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac 3000

tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag 3060

gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg 3120

gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta 3180

tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg 3240

ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata 3300

tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt 3360

ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc 3420

ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct 3480

tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa 3540

ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag 3600

tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc 3660

tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg 3720

actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca 3780

cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat 3840

gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg 3900

tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc 3960

ctgtcgggtt tcgccacctc tgacttgagc gtcgatttttt gtgatgctcg tcaggggggc 4020

ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc 4080

cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg 4140

cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga 4200

gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc 4260

attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa 4320

ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc 4380

gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg 4440

attacgccaa gctctcgaga tctag 4465

<210> 29

<211> 4405

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 29

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat tttagggtta gggttagggt tagggaaaaa actagtccac 1620

tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc 1680

gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agagagggac agatccgggc 1740

ccgcatgcgt cgacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg 1800

gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg 1860

aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc 1920

tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc 1980

tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg 2040

ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg 2100

tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa 2160

agggcctcgt gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga 2220

cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa 2280

tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt 2340

gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg 2400

cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag 2460

atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg 2520

agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg 2580

gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt 2640

ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga 2700

cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac 2760

ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc 2820

atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc 2880

gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac 2940

tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag 3000

gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg 3060

gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta 3120

tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg 3180

ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata 3240

tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt 3300

ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc 3360

ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct 3420

tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa 3480

ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag 3540

tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc 3600

tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg 3660

actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca 3720

cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat 3780

gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg 3840

tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc 3900

ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc 3960

ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc 4020

cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg 4080

cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga 4140

gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc 4200

attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa 4260

ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc 4320

gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg 4380

attacgccaa gctctcgaga tctag 4405

<210> 30

<211> 4465

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 30

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggtt tagggttagg gttagggtta gggaaaaatt tagggttagg gttagggtta 240

gggaaaaatt tagggttagg gttagggtta gggaaaaacc taggcatatg ccaagtacgc 300

cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct 360

tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga 420

tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa 480

gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc 540

caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg 600

aggtctatat aagcagagct cgtttagtga accgtcagat cgcctggaga cgccatccgg 660

actctaaggt aaatataaaa ttttttaagtg tataatgtgt taaactactg attctaattg 720

tttctctctt ttagattcca acctttggaa ctgaattccg cgggcccggg atccaccggt 780

cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga 840

gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc 900

cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg 960

gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca 1020

catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac 1080

catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga 1140

caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct 1200

ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca 1260

gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca 1320

gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga 1380

caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca 1440

catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta 1500

caagtaaagc ggccgctagg cctcacctgc gatctcgatg ctttatttgt gaaatttgtg 1560

atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt 1620

gcattcattt tatgtttcag gttcagggggg aggtgtggga ggttttttaa actagtccac 1680

tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc 1740

gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agagagggac agatccgggc 1800

ccgcatgcgt cgacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg 1860

gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg 1920

aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc 1980

tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc 2040

tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg 2100

ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg 2160

tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa 2220

agggcctcgt gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga 2280

cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa 2340

tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt 2400

gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg 2460

cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag 2520

atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg 2580

agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg 2640

gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt 2700

ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga 2760

cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac 2820

ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc 2880

atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc 2940

gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac 3000

tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag 3060

gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg 3120

gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta 3180

tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg 3240

ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata 3300

tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt 3360

ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc 3420

ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct 3480

tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa 3540

ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag 3600

tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc 3660

tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg 3720

actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca 3780

cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat 3840

gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg 3900

tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc 3960

ctgtcgggtt tcgccacctc tgacttgagc gtcgatttttt gtgatgctcg tcaggggggc 4020

ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc 4080

cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg 4140

cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga 4200

gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc 4260

attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa 4320

ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc 4380

gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg 4440

attacgccaa gctctcgaga tctag 4465

<210> 31

<211> 4435

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 31

aaagcttccc ggggggatct gggccactcc ctctctgcgc gctcgctcgc tcactgaggc 60

cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg 120

agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctggaggg gtggagtcgt 180

gacctaggca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 240

ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 300

tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 360

ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 420

ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 480

tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc 540

agatcgcctg gagacgccat ccggactcta aggtaaatat aaaattttta agtgtataat 600

gtgttaaact actgattcta attgtttctc tcttttagat tccaaccttt ggaactgaat 660

tccgcgggcc cgggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac 720

cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 780

gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac 840

caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca 900

gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 960

cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg 1020

cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1080

cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa 1140

cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca 1200

caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1260

cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa 1320

agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1380

cactctcggc atggacgagc tgtacaagta aagcggccgc taggcctcac ctgcgatctc 1440

gatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca 1500

ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt 1560

gggaggtttt ttaaatcgat gctagatgtt agcgtaaaaa gctagatgtt agcgtaaaaa 1620

gctagatgtt agcgtaaaaa actagtccac tccctctctg cgcgctcgct cgctcactga 1680

ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga 1740

gcgagcgcgc agagagggac agatccgggc ccgcatgcgt cgacaattca ctggccgtcg 1800

ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac 1860

atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac 1920

agttgcgcag cctgaatggc gaatggcgcc tgatgcggta ttttctcctt acgcatctgt 1980

gcggtatttc acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt 2040

taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc 2100

cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt 2160

caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta ttttttatagg 2220

ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc 2280

gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac 2340

aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt 2400

tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag 2460

aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg 2520

aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa 2580

tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc 2640

aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag 2700

tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa 2760

ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc 2820

taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg 2880

agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa 2940

caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa 3000

tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg 3060

gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag 3120

cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg 3180

caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt 3240

ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt 3300

aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac 3360

gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag 3420

atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg 3480

tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca 3540

gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga 3600

actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca 3660

gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc 3720

agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca 3780

ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa 3840

aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc 3900

cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc 3960

gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg 4020

cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat 4080

cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca 4140

gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca 4200

aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg 4260

actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac 4320

cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac 4380

aatttcacac aggaaacagc tatgaccatg attacgccaa gctctcgaga tctag 4435

<210> 32

<211> 6940

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 32

cagcagctgc gcgctcgctc gctcactgag gccgccccggg caaagcccgg gcgtcgggcg 60

acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120

atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180

tgctctagcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat gagcttggcc 240

cattgcatac gttgtatcca tatcataata tgtacattta tattggctca tgtccaacat 300

taccgccatg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat 360

tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg 420

gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa 480

cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact 540

tggcagtaca tcaagtgtat catatgccaa gtacgcccccc tattgacgtc aatgacggta 600

aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt 660

acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg 720

ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg 780

ggagttttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc 840

cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt 900

tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac 960

accgggaccg atccagcctc ccctcgaagc tgatcctgag aacttcaggg tgagtctatg 1020

ggacccttga tgttttcttt ccccttcttt tctatggtta agttcatgtc ataggaaggg 1080

gagaagtaac agggtacaca tattgaccaa atcagggtaa ttttgcattt gtaattttaa 1140

aaaatgcttt cttcttttaa tatacttttt tgtttatctt atttctaata ctttccctaa 1200

tctctttctt tcagggcaat aatgatacaa tgtatcatgc ctctttgcac cattctaaag 1260

aataacagtg ataatttctg ggttaaggca atagcaatat ttctgcatat aaatatttct 1320

gcatataaat tgtaactgat gtaagaggtt tcatattgct aatagcagct acaatccagc 1380

taccattctg cttttatttt atggttggga taaggctgga ttattctgag tccaagctag 1440

gcccttttgc taatcatgtt catacctctt atcttcctcc cacagctcct gggcaacgtg 1500

ctggtctgtg tgctggccca tcactttggc aaagaattcc gcgggcccgg gatccaccgg 1560

tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 1620

agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 1680

ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 1740

ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 1800

acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 1860

ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 1920

acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 1980

tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 2040

agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 2100

agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 2160

acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 2220

acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 2280

acaagtaaag cggccgctct agaggatcca agcttatcga taccgtcgac ctcgagggcc 2340

cagatctaat tcaccccacc agtgcaggct gcctatcaga aagtggtggc tggtgtggct 2400

aatgccctgg cccacaagta tcactaagct cgctttcttg ctgtccaatt tctattaaag 2460

gttcctttgt tccctaagtc caactactaa actgggggat attatgaagg gccttgagca 2520

tctggattct gcctaataaa aaacatttat tttcattgca atgatgtatt taaattattt 2580

ctgaatattt tactaaaaag ggaatgtggg aggtcagtgc atttaaaaca taaagaaatg 2640

aagagctagt tcaaaccttg ggaaaataca ctatatctta aactccatga aagaaggtga 2700

ggctgcaaac agctaatgca cattggcaac agcccctgat gcctatgcct tattcatccc 2760

tcagaaaagg attcaagtag aggcttgatt tggaggttaa agttttgcta tgctgtattt 2820

tacattactt attgttttag ctgtcctcat gaatgtcttt tcactaccca tttgcttatc 2880

ctgcatctct cagccttgac tccactcagt tctcttgctt agagatacca cctttcccct 2940

gaagtgttcc ttccatgttt tacggcgaga tggtttctcc tcgcctggcc actcagcctt 3000

agttgtctct gttgtcttat agaggtctac ttgaagaagg aaaaacaggg ggcatggttt 3060

gactgtcctg tgagcccttc ttccctgcct cccccactca cagtgacccg gaatccctcg 3120

acatctagag catggctacg tagataagta gcatggcggg ttaatcatta actacaagga 3180

acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg 3240

gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc 3300

gcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca 3360

gcctgaatgg cgaatggaat tccagacgat tgagcgtcaa aatgtaggta tttccatgag 3420

cgtttttcct gttgcaatgg ctggcggtaa tattgttctg gatattacca gcaaggccga 3480

tagtttgagt tcttctactc aggcaagtga tgttattact aatcaaagaa gtattgcgac 3540

aacggttaat ttgcgtgatg gacagactct tttactcggt ggcctcactg attataaaaa 3600

cacttctcag gattctggcg taccgttcct gtctaaaatc cctttaatcg gcctcctgtt 3660

tagctcccgc tctgattcta acgaggaaag cacgttatac gtgctcgtca aagcaaccat 3720

agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 3780

ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 3840

ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 3900

ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg 3960

ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata 4020

gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt 4080

tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat 4140

ttaacgcgaa ttttaacaaa atattaacgt ttacaattta aatatttgct tatacaatct 4200

tcctgttttt ggggcttttc tgattatcaa ccggggtaca tatgattgac atgctagttt 4260

tacgattacc gttcatcgat tctcttgttt gctccagact ctcaggcaat gacctgatag 4320

cctttgtaga gacctctcaa aaatagctac cctctccggc atgaatttat cagctagaac 4380

ggttgaatat catattgatg gtgatttgac tgtctccggc ctttctcacc cgtttgaatc 4440

tttacctaca cattactcag gcattgcatt taaaatatat gagggttcta aaaattttta 4500

tccttgcgtt gaaataaagg cttctcccgc aaaagtatta cagggtcata atgtttttgg 4560

tacaaccgat ttagctttat gctctgaggc tttattgctt aattttgcta attctttgcc 4620

ttgcctgtat gatttattgg atgttggaat tcctgatgcg gtattttctc cttacgcatc 4680

tgtgcggtat ttcacaccgc atatggtgca ctctcagtac aatctgctct gatgccgcat 4740

agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc 4800

tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt 4860

tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat 4920

aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg 4980

tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga 5040

gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac 5100

atttccgtgt cgcccttatt ccctttttttg cggcattttg ccttcctgtt tttgctcacc 5160

cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca 5220

tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc 5280

caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg 5340

ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac 5400

cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca 5460

taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg 5520

agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac 5580

cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg 5640

caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat 5700

taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg 5760

ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg 5820

cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc 5880

aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc 5940

attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt 6000

tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt 6060

aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt 6120

gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag 6180

cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca 6240

gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca 6300

agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg 6360

ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg 6420

cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct 6480

acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga 6540

gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc 6600

ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg 6660

agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg 6720

cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt 6780

tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc 6840

gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac 6900

gcaaaccgcc tctccccgcg cgttggccga ttcattaatg 6940

<210> 33

<211> 7257

<212> DNA

<213> Artificial sequences

<220>

<223> Synthetic sequences

<400> 33

cagcagctgc gcgctcgctc gctcactgag gccgccccggg caaagcccgg gcgtcgggcg 60

acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc 120

atcactaggg gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca 180

tgctctagcg gcctcggcct ctgcataaat aaaaaaaatt agtcagccat gagcttggcc 240

cattgcatac gttgtatcca tatcataata tgtacattta tattggctca tgtccaacat 300

taccgccatg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat 360

tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg 420

gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa 480

cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact 540

tggcagtaca tcaagtgtat catatgccaa gtacgcccccc tattgacgtc aatgacggta 600

aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt 660

acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg 720

ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg 780

ggagttttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc 840

cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctcgtt 900

tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac 960

accgggaccg atccagcctc ccctcgaagc tgatcctgag aacttcaggg tgagtctatg 1020

ggacccttga tgttttcttt ccccttcttt tctatggtta agttcatgtc ataggaaggg 1080

gagaagtaac agggtacaca tattgaccaa atcagggtaa ttttgcattt gtaattttaa 1140

aaaatgcttt cttcttttaa tatacttttt tgtttatctt atttctaata ctttccctaa 1200

tctctttctt tcagggcaat aatgatacaa tgtatcatgc ctctttgcac cattctaaag 1260

aataacagtg ataatttctg ggttaaggca atagcaatat ttctgcatat aaatatttct 1320

gcatataaat tgtaactgat gtaagaggtt tcatattgct aatagcagct acaatccagc 1380

taccattctg cttttatttt atggttggga taaggctgga ttattctgag tccaagctag 1440

gcccttttgc taatcatgtt catacctctt atcttcctcc cacagctcct gggcaacgtg 1500

ctggtctgtg tgctggccca tcactttggc aaagaattcc gcgggcccgg gatccaccgg 1560

tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 1620

agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 1680

ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 1740

ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 1800

acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 1860

ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 1920

acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 1980

tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 2040

agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 2100

agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 2160

acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 2220

acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 2280

acaagtaaag cggccgctct agaggatcca agcttatcga taccgtcgac ctcgagggcc 2340

cagatctaat tcaccccacc agtgcaggct gcctatcaga aagtggtggc tggtgtggct 2400

aatgccctgg cccacaagta tcactaagct cgctttcttg ctgtccaatt tctattaaag 2460

gttcctttgt tccctaagtc caactactaa actgggggat attatgaagg gccttgagca 2520

tctggattct gcctaataaa aaacatttat tttcattgca atgatgtatt taaattattt 2580

ctgaatattt tactaaaaag ggaatgtggg aggtcagtgc atttaaaaca taaagaaatg 2640

aagagctagt tcaaaccttg ggaaaataca ctatatctta aactccatga aagaaggtga 2700

ggctgcaaac agctaatgca cattggcaac agcccctgat gcctatgcct tattcatccc 2760

tcagaaaagg attcaagtag aggcttgatt tggaggttaa agttttgcta tgctgtattt 2820

tacattactt attgttttag ctgtcctcat gaatgtcttt tcactaccca tttgcttatc 2880

ctgcatctct cagccttgac tccactcagt tctcttgctt agagatacca cctttcccct 2940

gaagtgttcc ttccatgttt tacggcgaga tggtttctcc tcgcctggcc actcagcctt 3000

agttgtctct gttgtcttat agaggtctac ttgaagaagg aaaaacaggg ggcatggttt 3060

gactgtcctg tgagcccttc ttccctgcct cccccactca cagtgacccg gaatccctcg 3120

acaggtacct ttagggttag ggttagggtt agggaaaaat ttagggttag ggttagggtt 3180

agggaaaaat ttagggttag ggttagggtt agggaaaaat ttagggttag ggttagggtt 3240

agggaaaaat ttagggttag ggttagggtt agggaaaaac cctaacccta accctaaccc 3300

taaatttttt ccctaaccct aaccctaacc ctaaattttt tccctaaccc taaccctaac 3360

cctaaatttt ttccctaacc ctaaccctaa ccctaaattt tttccctaac cctaacccta 3420

accctaaatt ttttgctagc tctagagcat ggctacgtag ataagtagca tggcgggtta 3480

atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc 3540

tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc 3600

tcagtgagcg agcgagcgcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc 3660

ttcccaacag ttgcgcagcc tgaatggcga atggaattcc agacgattga gcgtcaaaat 3720

gtaggtattt ccatgagcgt ttttcctgtt gcaatggctg gcggtaatat tgttctggat 3780

attaccagca aggccgatag tttgagttct tctactcagg caagtgatgt tattactaat 3840

caaagaagta ttgcgacaac ggttaatttg cgtgatggac agactctttt actcggtggc 3900

ctcactgatt ataaaaacac ttctcaggat tctggcgtac cgttcctgtc taaaatccct 3960

ttaatcggcc tcctgtttag ctcccgctct gattctaacg aggaaagcac gttatacgtg 4020

ctcgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt 4080

ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt 4140

cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct 4200

ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg 4260

tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga 4320

gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc 4380

ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga 4440

gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caatttaaat 4500

atttgcttat acaatcttcc tgtttttggg gcttttctga ttatcaaccg gggtacatat 4560

gattgacatg ctagttttac gattaccgtt catcgattct cttgtttgct ccagactctc 4620

aggcaatgac ctgatagcct ttgtagagac ctctcaaaaa tagctaccct ctccggcatg 4680

aatttatcag ctagaacggt tgaatatcat attgatggtg atttgactgt ctccggcctt 4740

tctcacccgt ttgaatcttt acctacacat tactcaggca ttgcatttaa aatatatgag 4800

ggttctaaaa atttttatcc ttgcgttgaa ataaaggctt ctcccgcaaa agtattacag 4860

ggtcataatg tttttggtac aaccgattta gctttatgct ctgaggcttt attgcttaat 4920

tttgctaatt ctttgccttg cctgtatgat ttattggatg ttggaattcc tgatgcggta 4980

ttttctcctt acgcatctgt gcggtatttc acaccgcata tggtgcactc tcagtacaat 5040

ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc 5100

ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag 5160

ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt 5220

gatacgccta ttttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg 5280

cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa 5340

tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa 5400

gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct 5460

tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg 5520

tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg 5580

ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt 5640

atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga 5700

cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga 5760

attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac 5820

gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg 5880

ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac 5940

gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct 6000

agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct 6060

gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg 6120

gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat 6180

ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg 6240

tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat 6300

tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct 6360

catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa 6420

gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa 6480

aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc 6540

gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta 6600

gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct 6660

gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg 6720

atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag 6780

cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc 6840

cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg 6900

agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt 6960

tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg 7020

gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca 7080

catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg 7140

agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc 7200

ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatg 7257

<210> 34

<211> 15

<212> DNA

<213> Artificial sequences

<220>

<223> synthetic sequence; negative control

<400> 34

gctagatgtt agcgt 15

Claims (24)

1. A nucleic acid molecule comprising: A viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation. 2. A recombinant virus for delivering a desired function to mammalian cells, comprising: A viral genome covalently linked to inhibitory nucleic acid sequences that bind to TLR9 but do not trigger TLR9 activation. 3. The recombinant virus of claim 2, wherein the inhibitory nucleic acid sequence comprises the c41 oligonucleotide sequence TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1). 4. The recombinant virus of claim 2, wherein the inhibitory nucleic acid sequence is selected from the group consisting of: ODN 2088: TCC TGG CGG GGA AGT (SEQ ID NO: 2); ODN 4084-F: CCTGGATGGGAA (SEQ ID NO: 3); ODN INH-1: CCTGGATGGGAATTCCCATCCAGG (SEQ ID NO: 4); ODN INH-18: CCT GGA TGG GAA CTT ACC GCT GCA (SEQ ID NO: 5); ODN TTAGGG:TT AGG GTT AGG GTT AGG GTT AGG G (SEQ ID NO: 6); and G-ODN: CTC CTA TTG GGG GTT TCC TAT (SEQ ID NO: 7). 5. The recombinant virus of claim 2, wherein the inhibitory nucleic acid sequence is selected from the group consisting of: ODN 2114: TCCTGGAGGGGAAGT (SEQ ID NO: 16); ODN 4024: TCCTGGATGGGAAGT (SEQ ID NO: 17); ODN INH-4: TTCCCATCCAGGCCTGGATGGGAA (SEQ ID NO: 18); ODN INH-13: CTTACCGCTGCACCTGGATGGGAA (SEQ ID NO: 19); ODN Poly-G: GGGGGGGGGGGGGGGGGGGG (SEQ ID NO: 20); ODN GpG: TGACTGTGAAGGTTAGAGATGA (SEQ ID NO: 21); ODNIRS-869: TCCTGGAGGGGTTGT (SEQ ID NO: 22); ODN IRS-954:TGCTCCTGGAGGGGTTGT (SEQ ID NO:23); and ODN 21158: CCTGGCGGGG (SEQ ID NO: 24). 6. The recombinant virus of claim 4, wherein the inhibitory nucleic acid sequence is ODN TTAGGG (SEQ ID NO: 6). 7. A method of treating a mammal comprising: The recombinant virus of claim 2 is administered to a mammal in need thereof. 8. The method of claim 7, wherein the viral genome is an adeno-associated virus (AAV) genome. 9. The method of claim 7, wherein the inhibitory nucleic acid sequence comprises a c41 oligonucleotide sequence (SEQ ID NO: 1). 10. A method for preparing a viral genome of a recombinant virus, comprising: An inhibitory nucleic acid sequence is inserted into the viral genome, wherein the inhibitory nucleic acid sequence binds to TLR9 but does not trigger TLR9 activation. 11. The method of claim 10, further comprising: The viral genome is packaged in virions. 12. The method of claim 10, wherein the inhibitory nucleic acid sequence comprises a c41 oligonucleotide sequence (SEQ ID NO: 1). 13. A nucleic acid molecule comprising: Inverted terminal repeats (ITRs) and nucleic acid sequences that inhibit TLR9-mediated inflammation. 14. A recombinant virus for delivering a desired function to mammalian cells, comprising: Viral genomes comprising nucleic acid sequences that inhibit TLR9-mediated inflammation. 15. A method of preparing a viral genome of a recombinant virus, comprising: A nucleic acid sequence is inserted into the viral genome, wherein the nucleic acid sequence inhibits TLR9-mediated inflammation. 16. A nucleic acid vector comprising at least one nucleic acid sequence capable of inhibiting TLR9-mediated inflammation. 17. A method of reducing the immunogenicity of a modified virus having a genome, the method comprising inserting into the genome a nucleic acid sequence, the inhibitory nucleic acid sequence inhibits TLR9-mediated inflammation, whereby the The modified virus elicits a reduced inflammatory response in the host compared to a virus that does not contain the nucleic acid sequence. 18. A method of increasing the expression of a transgene introduced by a virus in a host cell, the method comprising introducing into the host a modified virus having a genome comprising a nucleic acid sequence that inhibits TLR9 mediation inflammation, whereby the modified virus results in higher expression of the transgene in the host cell compared to a virus that does not contain the nucleic acid sequence. 19. A composition comprising a viral capsid encapsulating a nucleic acid sequence that inhibits TLR9-mediated inflammation. 20. The recombinant virus of claim 2, wherein the inhibitory nucleic acid sequence is 95% identical to the c41 oligonucleotide sequence TGGCGCGCACCCACGGCCTG (SEQ ID NO: 1). 21. The recombinant virus of claim 2, wherein the inhibitory nucleic acid sequence is 95% identical to a sequence selected from the group consisting of: ODN 2088: TCC TGG CGG GGA AGT (SEQ ID NO: 2); ODN 4084-F: CCTGGATGGGAA (SEQ ID NO: 3); ODN INH-1: CCTGGATGGGAATTCCCATCCAGG (SEQ ID NO: 4); ODN INH-18: CCT GGA TGG GAA CTT ACC GCT GCA (SEQ ID NO: 5); ODN TTAGGG:TT AGG GTT AGG GTT AGG GTT AGG G (SEQ ID NO: 6); and G-ODN: CTC CTA TTG GGG GTT TCC TAT (SEQ ID NO: 7). 22. The recombinant virus of claim 2, wherein the inhibitory nucleic acid sequence is 95% identical to a sequence selected from the group consisting of: ODN 2114: TCCTGGAGGGGAAGT (SEQ ID NO: 16); ODN 4024: TCCTGGATGGGAAGT (SEQ ID NO: 17); ODN INH-4: TTCCCATCCAGGCCTGGATGGGAA (SEQ ID NO: 18); ODN INH-13: CTTACCGCTGCACCTGGATGGGAA (SEQ ID NO: 19); ODN Poly-G: GGGGGGGGGGGGGGGGGGGG (SEQ ID NO: 20); ODN GpG: TGACTGTGAAGGTTAGAGATGA (SEQ ID NO: 21); ODN IRS-869: TCCTGGAGGGGTTGT (SEQ ID NO: 22); ODN IRS-954:TGCTCCTGGAGGGGTTGT (SEQ ID NO:23); and ODN 21158: CCTGGCGGGG (SEQ ID NO: 24). 23. The method of claim 7, wherein the inhibitory nucleic acid sequence comprises SEQ ID NO:6 or SEQ ID NO:9. 24. The method of claim 7, wherein the inhibitory nucleic acid sequence comprises multiple copies of SEQ ID NO:6 and/or SEQ ID NO:9.

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